Document text
Principal Investigator: Mortimer Poncz
Organization: VERSITI BLOOD HEALTH, INC.
Fiscal Year: 2024
Award: $650,240
Funding agency: National Heart Lung and Blood Institute
ABSTRACT
Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a complication of adenoviral (AdV)-based
COVID-19 vaccination. Antibodies (Abs) against the 70-aa chemokine, platelet (Plt) factor 4 (PF4), have been
noted in both VITT and in the related immune, prothrombotic disorder, heparin-induced thrombocytopenia (HIT),
but target different sites on the human (h) PF4 tetramer. The site on hPF4 targeted in VITT is conserved in the
related chemokine, 94-aa -granule, Plt-basic protein (PBP) and its N-terminal-truncated isoforms (PBPi),
including 70-aa neutrophil-activating peptide 2 (NAP2) that uniquely activates neutrophils (PMNs) via CXCR2.
We hypothesize that PF4 and/or PBP can initiate VITT and contribute to its prothrombotic state. We show that
VITT Abs bind hPBPi and activate Plts. Mice injected with VITT plasma develop a prothrombotic state even in
the absence of PF4, and unlike in murine HIT, PMNs become incorporated into arterial as well as venular
thrombi. We also show by dynamic light scattering (DLS) that both chemokines bind directly to AdVs. In a murine
VITT model, Abs develop to either PF4 or PBPi. We will pursue our observation and test the above hypothesis
as follows: Aim 1: Characterize the range of antigenic targets of VITT Abs. We will confirm that VITT Abs,
VITT monoclonal (mo) Abs, and murine Abs generated in Aim 3 bind to hPBPi. We will also determine where
VITT Abs bind on hPBPi using select amino acid substitutions of hPBP. Aim 2: Examine the importance of
hPF4 and hPBPi in VITT in vitro and in vivo. We will test the relative contribution of hPF4 vs. hPBPi to
thrombus formation in vitro using an injured-endothelium microfluidic model. Transgenic FcRIIA+ mice that
express either no PF4 or PBP, or have the murine or human versions, will be injected with VITT Abs to recreate
the prothrombotic state to validate the importance of PF4 vs. PBP in the development of thrombi in vivo. Aim 3:
Examine the mechanistic basis for the onset of VITT in a murine model. We have developed a novel murine
VITT model that suggests that the prothrombotic state occurs after AdV vaccination independent of expressing
the COVID-19 spike protein. Both anti-PF4 and anti-PBPi Abs were noted. These studies were supported by
DLS studies which showed that both chemokines complex with AdV, but that some AdV bind better to hPF4 and
others to hPBPi. The mechanistic basis of VITT will be further pursued, including defining the site(s) by which
these chemokines interact with AdV, and the characterization of the formed Abs. Taken together, Project 3
should provide important, novel conceptual insights into VITT. A number of new VITT-like pro-thrombotic
disorders are being recognized, and our studies should also be applicable to these disorders. Finally, AdV-based
vaccines have been developed for other disorders, and our models in Aim 3 may guide AdV modifications to
enhance vaccine safety. These advances will greatly benefit from synergy between Dr. Poncz and the expertise
provided by shared Core B at Versiti's Blood Research Institute, as well as with the other Project Leaders in our
Program – each studying thematically related, clinically important immune thrombocytopenic disorders.
Terms: <2019-nCoV S protein><2019-nCoV spike glycoprotein><2019-nCoV spike protein><7S Gamma Globulin><AZD1222><Adenoviridae><Adenoviruses><Adjuvant><Alanine><Amino Acid Sequence><Amino Acid Substitution><Amino Acids><Antibodies><Antiheparin Factor><AstraZeneca COVID-19 vaccine><AstraZeneca coronavirus disease 2019 vaccine><Autoimmune Thrombocytopenias><Autoimmune Thrombocytopenic Purpura><Beta-TG><Binding><Binding Sites><Blood><Blood Neutrophil><Blood Plasma><Blood Platelet Factor IV><Blood Platelets><Blood Polymorphonuclear Neutrophil><Blood Reticuloendothelial System><Blood platelet factor 4><CDw128b><CDw128b Antigens><CMKAR2><COVID-19 S protein><COVID-19 infection><COVID-19 spike><COVID-19 spike glycoprotein><COVID-19 spike protein><COVID-19 vaccination><COVID-19 virus infection><COVID19 infection><CTAP III><CTAP3><CTAPIII><CXC Chemokine Ligand 7><CXC Chemokine Receptor 2><CXCL7><CXCR2><CXCR2 Protein><CXCR2 Receptors><Carotid Arteries><Cessation of life><Chemokine (C-X-C motif) Ligand 4><Chemokine (C-X-C) Receptor 2><Chemokine Receptor Gene><Chemotactic Cytokines><Clinical><Clinical Treatment Moab><Clotting><Coagulation><Coagulation Process><Combining Site><Complex><Complication><Connective Tissue-Activating Peptide III><Cytoplasmic Granules><Data><Deaminase><Death><Development><Diagnosis><Diagnostic><Disease><Disorder><ELISA><Endothelium><Enzyme-Linked Immunosorbent Assay><Factor 4><Femoral vein><GRO/MGSA Receptor><Genetics-Mutagenesis><Heparin><Heparin Binding><Heparin Neutralizing Protein><Heparinic Acid><High Affinity Interleukin 8 Receptor Type B><Homologous Chemotactic Cytokines><Human><IL-8RB><IL-8Rbeta><IL8 Receptor Type 2><IL8R2><IL8RB><IL8RB gene><Idiopathic Thrombocytopenic Purpura><IgG><Immune><Immune thrombocytopenia><Immunes><Immunobiology><Immunoglobulin G><Immunophysiology><In Vitro><Intercrines><Interleukin 8 Receptor Beta><Interleukin 8 Receptor Type 2><Interleukin-8 Receptor Type B><Interleukin-8 Receptors B><Interleukin-8B Receptor><Intramuscular><Isoforms><KO mice><Knock-out><Knock-out Mice><Knockout><Knockout Mice><LA-PF4><Laser injury><Life><Marrow Neutrophil><Marrow platelet><Medicine><Mice><Mice Mammals><Microfluidics><Modeling><Modern Man><Modernization><Modification><Molecular><Molecular Interaction><Monoclonal Antibodies><Murine><Mus><Mutagenesis><Mutagenesis Molecular Biology><N-terminal><NAP-2><NAP2><NH2-terminal><Nature><Neutrophil-Activating Peptide 2><Neutrophilic Granulocyte><Neutrophilic Leukocyte><Null Mouse><Outcome><PBP gene><PBP protein><PF4 Gene><PPBP><PPBP gene><Patients><Persons><Plasma><Plasma Enhancement><Plasma Serum><Platelet Basic Protein><Platelet Factor 4><Platelets><Polymorphonuclear Cell><Polymorphonuclear Leukocytes><Polymorphonuclear Neutrophils><Primary Protein Structure><Pro-Platelet Basic Protein><Protein Isoforms><Proteins><Publishing><Reaction><Reactive Site><Recombinant Platelet Factor 4><Research><Research Institute><Reticuloendothelial System, Serum, Plasma><Reverse engineering><Risk><Role><Rose Bengal><SARS-CoV-2 S><SARS-CoV-2 S protein><SARS-CoV-2 infection><SARS-CoV-2 spike><SARS-CoV-2 spike glycoprotein><SARS-CoV-2 spike protein><SARS-CoV-2 vaccination><SARS-CoV2 infection><SCYB4><SCYB7><SIS cytokines><Safety><Scanning><Severe acute respiratory syndrome coronavirus 2 S protein><Severe acute respiratory syndrome coronavirus 2 infection><Severe acute respiratory syndrome coronavirus 2 spike glycoprotein><Severe acute respiratory syndrome coronavirus 2 spike protein><Severe acute respiratory syndrome coronavirus 2 vaccination><Site><Small Inducible Cytokine B4><Small Inducible Cytokine Subfamily B, Member 4><Small Inducible Cytokine Subfamily B, Member 7><Spinal Column><Spine><Structure><TC1><TC2><TGB1><Testing><Therapeutic><Thrombocidin 1><Thrombocidin 2><Thrombocytes><Thrombocytopenia><Thromboglobulin, Beta-1><Thrombopenia><Thrombosis><Thrombus><Time><Transgenic Organisms><Treatment Period><Vaccination><Vaccinee><Vaccines><Venous><Vertebral column><Werlhof's Disease><aminoacid><arteriole><backbone><beta-Thromboglobulin><chemoattractant cytokine><chemokine><chemokine receptor><coronavirus disease 2019 S protein><coronavirus disease 2019 infection><coronavirus disease 2019 spike glycoprotein><coronavirus disease 2019 spike protein><coronavirus disease 2019 vaccination><develop a vaccine><develop vaccines><development of a vaccine><developmental><enzyme linked immunoassay><extracellular><gamma-Thromboglobulin><granule><heparin bound><heparin-induced thrombocytopenia><immune thrombocytopenic purpura><in vivo><infected with COVID-19><infected with COVID19><infected with SARS-CoV-2><infected with SARS-CoV2><infected with coronavirus disease 2019><infected with severe acute respiratory syndrome coronavirus 2><injured><insight><light scattering><mAbs><monoclonal Abs><mouse model><murine model><neutrophil><new vaccines><next generation vaccines><novel><novel vaccines><optimal therapies><optimal treatments><outcome following vaccination><outcome following vaccine><platelet factor IV><programs><protein sequence><result following vaccination><result following vaccine><social role><spike proteins on SARS-CoV-2><synergism><thromboembolic complications><thrombosis complications><thrombotic><thrombotic complications><thrombotic disease><thrombotic disorder><transfusion medicine><transgenic><treatment days><treatment duration><vaccinate against COVID-19><vaccinate against SARS-CoV-2><vaccinate against coronavirus disease 2019><vaccinate against severe acute respiratory syndrome coronavirus 2><vaccinated individual><vaccinated participant><vaccinated patient><vaccinated person><vaccinated subject><vaccination against COVID-19><vaccination against SARS-CoV-2><vaccination against Severe acute respiratory syndrome coronavirus 2><vaccination against coronavirus disease 2019><vaccination outcome><vaccination result><vaccine development><vaccine outcome><vaccine result><vaccine safety><vaccine strategy><venule><µfluidic><β-TG><β-Thromboglobulin>