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Principal Investigator: Mohammad Mohseni Sajadi
Organization: BALTIMORE VA MEDICAL CENTER
Fiscal Year: 2022
Funding agency: Veterans Affairs
Background/Rationale: As the Coronavirus Disease 2019 (COVID-19) epidemic expands across the United
States and the world, there are no proven therapies and little information is known regarding on immunity to
this virus. At this stage of the pandemic, all prevention and treatment strategies that show promise must be
explored. This proposal will focus on the polyclonal and monoclonal antibody responses to the severe acute
respiratory syndrome coronavirus-2 (SARS-CoV-2) envelope.
Objectives: The overarching goal of this program is to obtain a comprehensive understanding of the
polyclonal and monoclonal response to the SARS-CoV-2 envelope E, M, and S proteins. The specific aims of
this proposal are: 1) Deconvolute the polyclonal antibody response against the viral envelope of SARS-CoV-2;
2) Isolate neutralizing and non-neutralizing monoclonal antibodies against various epitopes of spike protein (S),
envelope protein (E), and membrane glycoprotein (M) of SARS-CoV-2; and 3) Map the corresponding
epitope(s) of the monoclonal antibodies.
Methods: We will obtain paired, acute and convalescent, samples from 60 inpatients and outpatients with
COVID-19 (30 have already been enrolled from the VA Maryland Health Care System and the University of
Maryland Medical System). We will screen samples by binding, live and pseudovirus neutralization. In this
proposal, we also will have access to BSL3 facilities on this campus that run neutralization assay with live virus
by one of the world experts in coronaviruses (Matt Frieman, PhD). Donors will be ranked on the basis of both
neutralization potency and breadth, and binding. The top three donors in each of the inpatient and outpatient
groups will be chosen for further study (top anti-RBD neutralization and breath, top anti- receptor binding domain
(RBD)-depleted plasma neutralization and breath, and top spike binding titers with non-neutralizing plasma).
In these six individuals, the anti-envelope antibody will be affinity purified; and fractionated using free-
flow-electrophoresis. This technique can separate antibodies based on charge, which will lead to separation
based on targeted epitopes as well. Individual fractions will be tested by binding and neutralization; and
characteristic biochemical and functional signatures of antibodies targeting each epitope will be ascertained.
Fractions of interest (different for each donor depending if neutralization or binding is targeted) will be sent for
mass spectrometry. B cell libraries will also be made from the convalescent IgG and IgA memory B cell pools;
and they will be interrogated using three complementary techniques including - acute phase plasmablast
repertoire analysis, subtraction analysis, and antigen baiting to identify potential antibodies.
Finally, mass spectrometry will be used to rank antibody candidates. 40 monoclonal antibodies (mAbs)
will be made and again be screened by neutralization potency, breadth, and isoelectric point, with top neutralizing
and binding antibodies (matching the respective profiles of the fractions that were targeted) to be moved forward
for fine epitope analysis by X-ray crystallography.
Impact: If successful, this project will yield a substantial understanding of neutralizing/non-neutralizing
antibodies and epitopes in COVID-19 disease, providing mAbs that can be moved into animal/human testing.
This research has direct relevance to the health of Veterans as any monoclonal antibodies isolated can
potentially be used for treatment and/or prevention of COVID-19.
Terms: <2019 novel corona virus><2019 novel coronavirus><2019-nCoV><2019-nCoV S protein><2019-nCoV spike glycoprotein><2019-nCoV spike protein><7S Gamma Globulin><AIDS Virus><Acquired Immune Deficiency Syndrome Virus><Acquired Immunodeficiency Syndrome Virus><Acute><Affinity><American><Animals><Antibodies><Antibody Response><Antigenic Determinants><Antigens><Assay><Award><B blood cells><B cell><B cells><B-Cells><B-Lymphocytes><B-cell><BSL-3 facility><BSL3 facility><Binding><Binding Determinants><Binding Sites><Bioassay><Biochemical><Biologic Assays><Biological Assay><Blood Plasma><COVID crisis><COVID epidemic><COVID infected patient><COVID pandemic><COVID patient><COVID positive patient><COVID-19><COVID-19 S protein><COVID-19 antibody><COVID-19 crisis><COVID-19 epidemic><COVID-19 global health crisis><COVID-19 global pandemic><COVID-19 health crisis><COVID-19 infected patient><COVID-19 infection><COVID-19 pandemic><COVID-19 patient><COVID-19 positive patient><COVID-19 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Immunodeficiency Viruses><IgA><IgG><Immunity><Immunoglobulin A><Immunoglobulin G><Individual><Infection><Inpatients><Isoelectric Point><Knowledge><LAV-HTLV-III><Lead><Libraries><Lymphadenopathy-Associated Virus><M protein><Maps><Maryland><Mass Photometry/Spectrum Analysis><Mass Spectrometry><Mass Spectroscopy><Mass Spectrum><Mass Spectrum Analyses><Mass Spectrum Analysis><Medical><Membrane Glycoproteins><Memory B Cell><Memory B-Lymphocyte><Methods><Modern Man><Molecular Interaction><Monoclonal Antibodies><Nucleoproteins><Out-patients><Outpatients><Patients><Pb element><Persons><Ph.D.><PhD><Phase><Plasma><Plasma Serum><Plasmablast><Plasmids><Preventative strategy><Prevention strategy><Preventive strategy><Production><Proteins><Protocol><Protocols documentation><Reactive Site><Research><Reticuloendothelial System, Serum, Plasma><Running><SARS corona virus 2><SARS-CO-V2><SARS-COVID-2><SARS-CoV-2><SARS-CoV-2 S protein><SARS-CoV-2 antibody><SARS-CoV-2 epidemic><SARS-CoV-2 global health 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