COVID-19 vaccine
Inventors
Barbieri, Joseph T. • Przedpelski, Amanda • Johnson, Eric A. • Pellett, Sabine • Tepp, William H.
Assignees
Medical College of Wisconsin • Wisconsin Alumni Research Foundation
Publication Number
US-12344635-B2
Publication Date
2025-07-01
Expiration Date
2041-03-24
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Abstract
Provided herein are antigenic peptides comprising the SARS-COV-2 spike protein receptor binding domain (CRBD) polypeptide or portions thereof, linked to a non-catalytic, non-toxic tetanus toxin variant (i.e., a modified tetanus toxin or “MTT”) and vaccine compositions comprising the same. In addition, provided herein are methods for making and using CRBD-MTT fusion proteins as immunogenic agents.
Core Innovation
The invention provides antigenic peptides comprising the SARS-COV-2 spike protein receptor binding domain (CRBD) polypeptide or portions thereof, linked to a non-catalytic, non-toxic modified tetanus toxin variant (MTT), and vaccine compositions comprising the same. It also encompasses methods for making and using CRBD-MTT fusion proteins as immunogenic agents. These fusion proteins are designed to elicit an immune response against SARS-COV-2 by presenting the viral receptor binding domain in conjunction with a genetically modified tetanus toxin that has significantly reduced toxicity.
The problem being addressed is the urgent need for improved vaccination platforms against COVID-19, caused by SARS-COV-2. Current understanding identifies the spike protein and more specifically the receptor binding domain as critical in viral binding to host cells. Emerging structural insights of this domain facilitate the design of antigenic fusion proteins that could provide effective vaccination. The background highlights the lack of optimized vaccines that combine immunogenic viral epitopes with a safe and effective carrier or adjuvant platform.
The core innovation lies in fusing the SARS-COV-2 receptor binding domain, particularly residues 330-521 of the spike protein, to a genetically inactivated tetanus toxin protein that contains multiple mutations substantially reducing catalytic and receptor binding activities and toxicity. These MTTs include specific point mutations at key residues such as R372, Y375, E234, K768, R1226, and W1289, among others, which disable toxin functions. The fusion proteins may connect the CRBD and MTT either directly, via a linker polypeptide (e.g., poly-glycine), or by chemical cross-linking. This approach leverages a well-characterized and safe toxoid platform to present the SARS-COV-2 antigen, promoting immune recognition without the risks associated with active toxins.
Claims Coverage
The patent claims include multiple independent claims covering the composition, encoding polynucleotides, vectors, methods of production, and methods of use. The main inventive features relate to the design of the fusion protein, its genetic encoding, its chemical crosslinking, and its application as a vaccine.
Fusion protein comprising mutated tetanus toxin and SARS-COV-2 receptor binding domain
A fusion protein comprising (i) a modified tetanus toxin (MTT) polypeptide with mutations at residues R372 and Y375 and at least two mutations among E234, K768, R1226, and W1289, resulting in reduced toxicity and receptor binding compared to wild type, and (ii) a SARS-COV-2 spike protein receptor binding domain (CRBD) polypeptide or a portion thereof.
Linker or chemical crosslink connection between fusion protein components
The MTT polypeptide and the CRBD polypeptide are connected either by a linker polypeptide, such as a poly-glycine sequence, or by chemical crosslinking using agents like formaldehyde, DSS, SMCC, Sulfo-EGS, BS3, or DSP.
Polynucleotides encoding the CRBD-MTT fusion protein
Nucleic acid sequences that encode the CRBD-MTT fusion protein with at least 95% sequence identity to specified sequences, enabling recombinant production of the fusion protein.
Vectors comprising polynucleotides encoding the fusion protein
Expression or recombinant vectors incorporating the polynucleotides encoding the CRBD-MTT fusion proteins for host cell transformation and protein production.
Methods for producing chemically crosslinked CRBD-MTT fusion protein
Processes involving obtaining polypeptide compositions of the MTT and CRBD components and chemically crosslinking them using crosslinking agents under conditions sufficient to produce the fusion protein.
Methods of reducing risk of COVID-19 by administering fusion protein
Therapeutic methods to induce immune responses by administering a therapeutically effective amount of the CRBD-MTT fusion protein to reduce the risk of developing COVID-19.
The claims comprehensively cover compositions of engineered fusion proteins with multiple defined mutations in tetanus toxin linked to the SARS-COV-2 receptor binding domain, nucleic acid sequences encoding them, means of producing such fusion proteins including chemical crosslinking, vectors for expression, and therapeutic methods of administration as vaccines to induce immunity against COVID-19.
Stated Advantages
The modified tetanus toxin fusion proteins have greatly reduced toxicity and receptor binding compared to wild-type tetanus toxin, enhancing safety for vaccine use.
The fusion proteins combine the immunogenic receptor binding domain of SARS-COV-2 with a widely studied and effective vaccine platform based on tetanus toxoid variants.
The CRBD-MTT fusion proteins are soluble when produced in E. coli, facilitating recombinant production and purification.
The platform permits chemical crosslinking or linker peptides for flexible fusion protein construction.
The use of genetically modified tetanus toxin obviates the need for toxic formalin detoxification, reducing production complexity and risk.
Documented Applications
Use as a vaccine composition to induce an immune response against SARS-COV-2 in a subject thereby reducing the risk of developing COVID-19.
Production of CRBD-MTT fusion proteins by recombinant expression in host cells such as E. coli for use as immunogenic agents.
Chemical crosslinking of separately produced CRBD and MTT polypeptides to form fusion proteins for vaccine formulations.
Administration of the fusion proteins to mammals to confer active immunization against SARS-COV-2.
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