ABSTRACT
The entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) into host cells requires binding of the viral spike glycoprotein to the angiotensin-converting enzyme 2 (ACE2) receptor, which triggers subsequent conformational changes to facilitate viral and cellular fusion at the plasma membrane or following endocytosis. Here, we experimentally identified selective and broad inhibitors of SARS-CoV-2 entry that share a tricyclic ring (or similar) structure. The inhibitory effect was restricted to early steps during infection and the entry inhibitors interacted with the receptor binding domain of the SARS-CoV-2 spike but did not significantly interfere with receptor (ACE2) binding. Instead, some of these compounds induced conformational changes or affected spike assembly and blocked SARS-CoV-2 spike cell-cell fusion activity. The broad inhibitors define a highly conserved binding pocket that is present on the spikes of SARS-CoV-1, SARS-CoV-2, and all circulating SARS-CoV-2 variants tested and block SARS-CoV spike activity required for mediating viral entry. These compounds provide new insights into the SARS-CoV-2 spike topography, as well as into critical steps on the entry pathway, and can serve as lead candidates for the development of broad-range entry inhibitors against SARS-CoVs.
Subject(s)
Angiotensin-Converting Enzyme 2 , COVID-19 , Glycoproteins , Humans , Peptidyl-Dipeptidase A/metabolism , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Virus InternalizationABSTRACT
We describe the production of single-cycle (sc) and replication-competent recombinant vesicular stomatitis viruses (rcVSVs) displaying heterologous envelope glycoproteins (Envs) on their surface. We prepare scVSVs by transiently expressing HIV-1 Envs or SARS-CoV-2 spike followed by infection of the cells with scVSV particles, which do not carry the vsv-g gene. To prepare rcVSVs, we replace the vsv-g with a specific env-encoding gene, transfect cells with multiple plasmids for production of the genomic RNA and viral proteins, and rescue replication-competent viruses.
Subject(s)
Recombinant Proteins , Spike Glycoprotein, Coronavirus , Vesicular Stomatitis/genetics , env Gene Products, Human Immunodeficiency Virus , Animals , COVID-19/virology , Cell Line , Cricetinae , HIV-1/genetics , Humans , Protein Engineering , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , env Gene Products, Human Immunodeficiency Virus/chemistry , env Gene Products, Human Immunodeficiency Virus/genetics , env Gene Products, Human Immunodeficiency Virus/metabolismABSTRACT
The COVID-19 (coronavirus disease 2019) pandemic has spread worldwide, leading to the deaths of millions and changing the way we live; we all hope to see the end of the pandemic soon. Nonetheless, an urgent need for medical interventions led to unprecedented and focused research efforts to translate scientific knowledge to new therapeutic and preventative interventions. Procedures were simplified, and new norms were established to expedite high-quality scientific output. We do hope that these changes will be adopted and streamlined to advance science in the future.