R01AI163019

Mechanism and Inhibition of SARS-CoV-2 Entry

The long-term goal of this study is to understand how Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causal agent of Coronavirus Disease 2019 (COVID-19), enters cells and how to block that process through the use of therapeutics.

Like other enveloped viruses, SARS-CoV-2 cell entry begins with engagement at the cell surface and is completed on release of the viral contents following membrane fusion. During the process of cell entry, the SARS-CoV-2 spike protein (S) engages the cellular receptor, Angiotensin Converting Enzyme (ACE2). Proteolytic activation of S is required to activate the fusion machinery, which can be achieved by cell surface or endosomal proteases. This poses a model of cell surface and endosomal entry routes that depend on engagement of different host-cell molecules that vary among cell types.

To interrogate the entry pathway of SARS-CoV-2, we developed a set of unique tools that permit application of single virion imaging approaches to track productive entry routes in an unbiased way and to help identify host factors coopted during viral entry. This imaging is facilitated by the use of a chimeric Vesicular Stomatitis Virus (VSV) in which its glycoprotein gene (G) was replaced with the spike (S) gene of SARS-CoV-2. Inhibition of VSV-SARS-CoV-2 infection with monoclonal antibodies, soluble receptor, and small molecule inhibitors correlates closely with inhibition of a clinical isolate of SARS-CoV-2, corroborating that the chimera is an effective BSL2 surrogate to study SARS-CoV-2 S-mediated entry.

This permits us to genetically modify a core protein of the VSV ribonucleoprotein core to render the particles visible by fluorescent microscopy. By combining this imaging approach with genetic, chemical, and biological perturbations, we will map the entry routes of VSV-SARS-CoV-2 and then examine the effect of those perturbations on infection of cells with a clinical isolate of SARS-CoV-2.

We will use this approach to determine how countermeasures currently in clinical trials, including monoclonal antibodies, soluble ACE2, and two small molecule inhibitors (Apilimod and Nafamostat), block entry. Using genome-wide loss-of-function screens, we will also interrogate the requirements for entry of SARS-CoV-2 under native and perturbed conditions to uncover new host proteins that are coopted during entry as potential additional targets for therapeutic intervention.

Successful completion of this work will define the entry pathways that lead to productive SARS-CoV-2 infection, inform the mechanism by which multiple molecules in clinical development interfere with that process, and unearth new host factors that are coopted during the entry pathway.

Washington University

TO ASSIST PUBLIC AND PRIVATE NONPROFIT INSTITUTIONS AND INDIVIDUALS TO ESTABLISH, EXPAND AND IMPROVE BIOMEDICAL RESEARCH AND RESEARCH TRAINING IN INFECTIOUS DISEASES AND RELATED AREAS, TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS. TO ASSIST PUBLIC, PRIVATE AND COMMERCIAL INSTITUTIONS TO CONDUCT DEVELOPMENTAL RESEARCH, TO PRODUCE AND TEST RESEARCH MATERIALS, TO PROVIDE RESEARCH SERVICES AS REQUIRED BY THE AGENCY FOR PROGRAMS IN INFECTIOUS DISEASES, AND CONTROLLING DISEASE CAUSED BY INFECTIOUS OR PARASITIC AGENTS, ALLERGIC AND IMMUNOLOGIC DISEASES AND RELATED AREAS. PROJECTS RANGE FROM STUDIES OF MICROBIAL PHYSIOLOGY AND ANTIGENIC STRUCTURE TO COLLABORATIVE TRIALS OF EXPERIMENTAL DRUGS AND VACCINES, MECHANISMS OF RESISTANCE TO ANTIBIOTICS AS WELL AS RESEARCH DEALING WITH EPIDEMIOLOGICAL OBSERVATIONS IN HOSPITALIZED PATIENTS OR COMMUNITY POPULATIONS AND PROGRESS IN ALLERGIC AND IMMUNOLOGIC DISEASES. BECAUSE OF THIS DUAL FOCUS, THE PROGRAM ENCOMPASSES BOTH BASIC RESEARCH AND CLINICAL RESEARCH. SMALL BUSINESS INNOVATION RESEARCH (SBIR) PROGRAM EXPANDS AND IMPROVES PRIVATE SECTOR PARTICIPATION IN BIOMEDICAL RESEARCH. THE SBIR PROGRAM INTENDS TO INCREASE AND FACILITATE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, TO INCREASE SMALL BUSINESS PARTICIPATION IN FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. THE SMALL BUSINESS TECHNOLOGY TRANSFER (STTR) PROGRAM STIMULATES AND FOSTERS SCIENTIFIC AND TECHNOLOGICAL INNOVATION THROUGH COOPERATIVE RESEARCH AND DEVELOPMENT CARRIED OUT BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO FOSTER TECHNOLOGY TRANSFER BETWEEN SMALL BUSINESS CONCERNS AND RESEARCH INSTITUTIONS, TO INCREASE PRIVATE SECTOR COMMERCIALIZATION OF INNOVATIONS DERIVED FROM FEDERAL RESEARCH AND DEVELOPMENT, AND TO FOSTER AND ENCOURAGE PARTICIPATION OF SOCIALLY AND ECONOMICALLY DISADVANTAGED SMALL BUSINESS CONCERNS AND WOMEN-OWNED SMALL BUSINESS CONCERNS IN TECHNOLOGICAL INNOVATION. RESEARCH CAREER DEVELOPMENT AWARDS SUPPORT THE DEVELOPMENT OF SCIENTISTS DURING THE FORMATIVE STAGES OF THEIR CAREERS. INDIVIDUAL NATIONAL RESEARCH SERVICE AWARDS (NRSAS) ARE MADE DIRECTLY TO APPROVE APPLICANTS FOR RESEARCH TRAINING IN SPECIFIED BIOMEDICAL SHORTAGE AREAS. IN ADDITION, INSTITUTIONAL NATIONAL RESEARCH SERVICE AWARDS ARE MADE TO ENABLE INSTITUTIONS TO SELECT AND MAKE AWARDS TO INDIVIDUALS TO RECEIVE TRAINING UNDER THE AEGIS OF THEIR INSTITUTIONAL PROGRAM.

93.855 - Allergy and Infectious Diseases Research

National Institute of Allergy and Infectious Diseases (NIAID) [HHS - NIH]

Missouri United States

State-Wide

NIH Research Project Grant (Parent R01 Clinical Trial Not Allowed) (PA-20-185)

Amendment Since initial award the total obligations have increased 393% from $766,721 to $3,776,105.