# US 11,130,787 B2 · Alphaherpesvirus glycoprotein d-encoding nucleic acid constructs and methods

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- Number: US 11,130,787 B2
- Document ID: US11130787B2
- Status: Granted
- Kind: B2
- Application number: 16/917,899
- Filed: 2020-06-30
- Published: 2020-10-15
- Granted: 2021-09-28
- Earliest claimed: 2020-06-11
- Assignees: MBF Therapeutics, Inc.
- Applicants: MBF Therapeutics, Inc.
- Inventors: Lorraine Keller; Malla Padidam
- Web page: https://patentgrep.com/patent/US11130787B2

## CPC

- C07K 14/005: from viruses
- A61K 39/385: Haptens or antigens, bound to carriers
- A61P 35/00: Antineoplastic agents
- C07K 14/70539: {MHC-molecules, e.g. HLA-molecules}
- A61K 2039/53: {DNA (RNA) vaccination}
- A61K 2039/552: {Veterinary vaccine}
- A61K 2039/572: {cytotoxic response}
- A61K 2039/575: {humoral response}
- A61K 2039/6075: {Viral proteins}
- A61K 2039/80: {Vaccine for a specifically defined cancer}
- C07K 2319/00: Fusion polypeptide
- C07K 2319/40: containing a tag for immunodetection, or an epitope for immunisation
- C07K 2319/70: containing domain for protein-protein interaction
- C12N 2710/10343: viral genome or elements thereof as genetic vector
- C12N 2710/12022: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2710/12034: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2710/16021: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2710/16622: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2710/16634: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2710/16722: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2710/16734: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2750/10022: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2760/16121: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2760/16122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2760/16134: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2770/10022: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2770/10034: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2770/20021: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2770/20022: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2770/20034: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

This invention provides new compositions comprising nucleotide sequence(s) encoding alphaherpesvirus glycoprotein D protein(s) (gDP(s)) and antigen(s) that induce immune responses. Such sequences typically encode gDP:antigen fusion proteins and typically also include feature(s) that significantly enhance immune responses such as (a) sequences encoding ITIC signal transducing adaptor proteins, e.g., SLAM-associated proteins (SAPs), Ewing's sarcoma-associated transcript 2 proteins, or both, or non-gDP checkpoint inhibitor(s); (b) sequences encoding antigen-associated targeting sequences, e.g., polyubiquitin sequences; (c) deimmunized/modified antigen-encoding sequences; (d) gDP(s) with modified sequence(s); (e) expression-enhancing introns; (f) transfection-facilitating agents; or (g) combinations thereof. Methods of using such constructs to induce immune responses and other methods and compositions also are provided, including methods of using such constructs in animals not known to express Herpesvirus entry mediator (HVEM) receptors (e.g., pigs), animals not under disease agent-associated checkpoint inhibition, and other contexts.

## Claims

### Claim 1

A composition comprising an effective amount of:
(A) a single DNA plasmid that comprises
(1) a nucleotide sequence that encodes
(a) a fusion protein, the fusion protein comprising
(i) one or more alphaherpesvirus glycoprotein D (gD) amino acid sequences comprising one or more gD receptor binding domains, and (ii) one or more antigens that are associated with a disease-causing agent, wherein the one or more gD amino acid sequences detectably facilitate dendritic cell uptake of the fusion protein, and
(b) one or more Ewing's Sarcoma-associated Transcript-2 (EAT-2) amino acid sequences, and
(2) one or more promoters that increase expression of the fusion protein and the one or more EAT-2 amino acid sequences;
(B) two or more DNA plasmids that collectively comprise
(1) nucleotide sequences that encode
(a) a fusion protein, the fusion protein comprising
(i) one or more gD amino acid sequences comprising one or more gD receptor binding domains and (ii) one or more antigens associated with a disease-causing agent, wherein the one or more gD sequences detectably facilitate dendritic cell uptake of the fusion protein, and
(b) one or more EAT-2 amino acid sequences, and
(2) one or more promoters that increase expression of the fusion protein and the one or more EAT-2 amino acid sequences;
(C) a single mRNA molecule that comprises a nucleotide sequence that encodes
(a) a fusion protein, the fusion protein comprising
(i) one or more gD amino acid sequences comprising one or more gD receptor binding domains, wherein the one or more gD sequences detectably facilitate dendritic cell uptake of the fusion protein, and (ii) one or more antigens associated with a disease-causing agent, and
(b) one or more EAT-2 amino acid sequences; or
(D) two or more mRNA molecules that collectively comprise nucleotide sequences that encode
(a) a fusion protein, the fusion comprising
(i) one or more gD amino acid sequences comprising one or more gD receptor binding domains, wherein the one or more gD sequences detectably facilitate dendritic cell uptake of the fusion protein, and (ii) one or more antigens associated with a disease-causing agent, and
(b) one or more EAT-2 amino acid sequences.

### Claim 2 (depends on 1)

The composition of claim 1, wherein at least one of the antigens are associated with one or more polyubiquitin sequences.

### Claim 3 (depends on 2)

The composition of claim 2, wherein at least one of the one or more gD sequences exhibits a detectable checkpoint inhibition function in target recipients.

### Claim 4 (depends on 3)

The composition of claim 3, wherein the one or more nucleotide sequences of the one or more DNA plasmids or one or more mRNA molecules further encode one or more non-gD checkpoint modulating amino acid sequences.

### Claim 5 (depends on 2)

The composition of claim 2, wherein the one or more gD sequences do not exhibit checkpoint inhibition in target recipients and the one or more nucleotide sequences of the one or more DNA plasmids or the one or more mRNA molecules further encode one or more non-gD checkpoint modulating amino acid sequences.

### Claim 6 (depends on 2)

The composition of claim 2, wherein at least one of the antigens encoded by the one or more nucleotide sequences of the one or more DNA plasmids or the one or more mRNA molecules comprise a mutation that eliminates a glycosylation site in the corresponding unmutated antigen.

### Claim 7 (depends on 1)

The composition of claim 1, wherein the composition comprises an effective amount of one or more DNA plasmids that are associated with transfection-facilitating calcium phosphate nanoparticles.

### Claim 8 (depends on 7)

The composition of claim 7, wherein the one or more plasmids further comprise one or more expression-enhancing introns.

### Claim 9 (depends on 1)

The composition of claim 1, wherein the one or more nucleotide sequences of the one or more DNA plasmids encode a plurality of antigens, wherein at least two of the antigens, when initially expressed, are separated from each other by an amino acid sequence comprising one or more self-cleavage sites.

### Claim 10 (depends on 7)

The composition of claim 7, wherein the one or more nucleotide sequences of the one or more DNA plasmids further comprise a triclosan selection marker.

### Claim 11 (depends on 8)

The composition of claim 8, wherein the one or more antigens comprise one or more pathogen antigens.

### Claim 12 (depends on 11)

The composition of claim 11, wherein the one or more pathogen antigens comprise one or more Porcine Circovirus antigens, one or more Porcine Reproductive and Respiratory Syndrome Virus antigens, one or more African Swine Fever Virus Antigens, or a combination of some or all thereof.