High throughput screening and drug discovery for antagonists of the Ebola VP40 protein assembly

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Principal Investigator: Ryan P Bennett
Organization: OYAGEN, INC.
Fiscal Year: 2024
Award: $298,000
Funding agency: National Institute of Allergy and Infectious Diseases

PROJECT SUMMARY
This R43 proposal answers the call of the RFA PA-22-176 for assay development and
chemical probe screening by addressing the unmet need for development of antiviral
treatments for Ebola patients through an innovative high-content small molecule screen
for antagonists of the Ebola VP40 matrix protein. There is a need for a fast-acting therapy
that is independent of the immune system and targets essential viral proteins. Such a
novel therapeutic is anticipated to enhance the survival probability for infected people in
hot zones of an Ebola outbreak. We chose to target EBOV VP40 because it is absolutely
required for EBOV particle assembly at the cell membrane, is capable of budding virus-
like particles (VLPs) when expressed in isolation and VP40 protein-protein interaction
domains have been structurally determined. Further guiding this application is our
published pilot screen which established proof-of-concept by demonstrating the
accessibility of Ebola VP40 protein-protein interactions to sangivamycin, a dual acting
small molecule antagonist of both EBOV VP40 assembly of virions and the viral
replication machinery. Given this success and due to anticipated complications with
efficacy and MOA studies inherent to compounds with dual targets, our Specific Aims
propose to screen a library of ~123,000 small molecule compounds to identify antagonists
of VP40 accumulation at the cell membrane for VLP formation and release from cells
through a fully automated, quantitative, and high-content assay. The assay has been
vetted to quantify the effect of small molecules on the cellular distribution of a fluorescent
VP40 expressed in 293T cells. Hits validated as dose-dependent by qHTS and displaying
low cytotoxicity will be further prioritized based on their absolute requirement for VP40 in
antiviral mechanism of action through counter screening with the VP40- independent
minigenome assay. Lead compounds also will be prioritized by their favorable ADMET
profiles. Our proposed critical path anticipates identifying 2-4 dose-dependent, VP40-
selective antagonists with potent antiviral activity that display low cytotoxicity for future
medicinal chemistry and preclinical development.

Terms: <Address><Anti-viral Agents><Area><Assay><BSL-4 facility><BSL4 facility><Back><Belgian Congo><Bioassay><Biogenesis><Biological Assay><Blood Plasma Cell><Cause of Death><Cell Body><Cell Cycle><Cell Division Cycle><Cell membrane><Cell-Mediated Cytolysis><Cell-Mediated Lympholysis><Cells><Cellular Cytotoxicity><Characteristics><Chemicals><Communities><Complex><Country><Critical Paths><Critical Pathways><Cytoplasm><Cytoplasmic Membrane><Democratic Republic of the Congo><Disease Outbreaks><Diversity Library><Dorsum><Dose><Drugs><Dryness><EBOV><Ebola><Ebola virus><Ebola-like Viruses><Evaluation><Filoviridae><Filovirus><Future><Gene Transcription><Gene variant><Genetic Transcription><History><Homo><Immune Targeting><Immune response><Immune system><Immunize><Immunological response><In Vitro><Individual><Infection><Intervention><Intervention Strategies><Lead><Libraries><Life><Link><Location><Lymphocyte Cytotoxicity><Lymphocytotoxicity><Lytotoxicity><Medication><Medicinal Chemistry><Membrane><Origin of Life><Outbreaks><Pathology><Patients><Pb element><Persons><Pharmaceutic Chemistry><Pharmaceutical Chemistry><Pharmaceutical Preparations><Plasma Cells><Plasma Membrane><Plasmacytes><Population><Powder dose form><Powders><Pro-Mega><Probability><Production><Promega><Property><Protein Binding Domain><Protein Binding Motif><Protein-Protein Interaction Domain><Proteins><Publishing><RNA Expression><Recording of previous events><Reporter><Research><Risk><Services><Structure><Symptoms><Testing><Texas><Therapeutic><Therapeutic Agents><Transcript><Transcription><Transfection><Travel><Triage><Vaccination><Viral><Viral Activity><Viral Function><Viral Gene Products><Viral Gene Proteins><Viral Genome><Viral Physiology><Viral Proteins><Virion><Virus><Virus Assembly><Virus Particle><Virus Replication><Virus-like particle><World Health Organization><ZEBOV><Zaire><Zaire Ebola virus><Zaire ebolavirus><allele variant><allelic variant><antagonism><antagonist><anti-viral compound><anti-viral development><anti-viral drug development><anti-viral drugs><anti-viral medication><anti-viral therapeutic><anti-viral therapeutic development><anti-viral therapy development><anti-virals><antiviral development><antiviral drug development><antiviral therapeutic development><antiviral therapy development><assay development><biosafety level 4 facility><cell mediated cytotoxicity><cell type><cytotoxic><cytotoxicity><developing anti-viral agent><developing anti-viral drug><developing anti-viral therapeutic><developing anti-viral therapy><developing antiviral agent><developing antiviral drug><developing antiviral therapeutic><developing antiviral therapy><drug candidate><drug discovery><drug/agent><effective therapy><effective treatment><emergent outbreak><emerging outbreak><extracellular vesicles><genetic variant><genomic variant><heavy metal Pb><heavy metal lead><high-throughput drug screening><histories><host response><imaging system><immune system response><immunoresponse><indexing><inhibitor><innovate><innovation><innovative><interventional strategy><medical countermeasure><membrane assembly><membrane structure><migration><mutant><new approaches><new drug treatments><new drugs><new outbreak><new pharmacological therapeutic><new therapeutic approach><new therapeutic intervention><new therapeutic strategies><new therapeutics><new therapy><new therapy approaches><new treatment approach><new treatment strategy><next generation therapeutics><novel><novel approaches><novel drug treatments><novel drugs><novel outbreak><novel pharmaco-therapeutic><novel pharmacological therapeutic><novel strategies><novel strategy><novel therapeutic approach><novel therapeutic intervention><novel therapeutic strategies><novel therapeutics><novel therapy><novel therapy approach><pandemic concern><pandemic potential><pandemic risk><pandemic threat><particle><past outbreak><plasmalemma><plasmocyte><pre-clinical development><preclinical development><previous outbreak><prior outbreak><protein crosslink><protein protein interaction><sangivamycin><screening><screenings><small molecular inhibitor><small molecule><small molecule inhibitor><social><success><targeted drug therapy><targeted drug treatments><targeted therapeutic><targeted therapeutic agents><targeted therapy><targeted treatment><viral assembly><viral multiplication><viral replication><virus genome><virus multiplication><virus protein><virus-like nanoparticles><viruslike particle>