# US 2021/0332084 A1 · Optimized zika virus envelope gene and expression thereof

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- Number: US 2021/0332084 A1
- Document ID: US20210332084A1
- Status: Published application
- Kind: A1
- Application number: 16/320,983
- Filed: 2017-07-26
- Published: 2021-10-28
- Earliest claimed: 2016-07-27
- Assignees: Hawaii Biotech, Inc.
- Applicants: Hawaii Biotech Inc.
- Inventors: David E. Clements
- Web page: https://patentgrep.com/patent/US20210332084A1

## CPC

- C07K 14/005: from viruses
- A61K 39/12: Viral antigens
- A61K 39/39: characterised by the immunostimulating additives, e.g. chemical adjuvants
- A61P 31/14: for RNA viruses
- C07K 14/18: Togaviridae; {Flaviviridae}
- C12N 15/85: for animal cells
- A61K 2039/55555: {Liposomes; Vesicles, e.g. nanoparticles; Spheres, e.g. nanospheres; Polymers}
- A61K 2039/55566: {Emulsions, e.g. Freund's adjuvant, MF59}
- A61K 2039/55577: {Saponins; Quil A; QS21; ISCOMS}
- C12N 2710/14143: viral genome or elements thereof as genetic vector
- C12N 2770/24122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2770/24134: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2770/24152: relating to complementing cells and packaging systems for producing virus or viral particles
- C12N 2770/24171: Demonstrated in vivo effect
- C12N 2800/22: Vectors comprising a coding region that has been codon optimised for expression in a respective host
- 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

The present invention is directed to the expression and secretion the Zika virus envelope protein. Elements of the pre-membrane and envelope sequence have been modified to enhance the expression of the envelope protein as a secreted product in the culture medium of transformed insect cell lines. The expressed and purified product is suitable as a vaccine antigen.

## Claims

### Claim at position 1

1-27. (canceled)

### Claim 28

An expression vector comprising a DNA sequence encoding a Zika virus pre-membrane and envelope protein, wherein expression of the DNA sequence results in secretion of a soluble envelope protein in the culture medium.

### Claim 29 (depends on 28)

The expression vector of claim 28, wherein the DNA sequence comprises SEQ ID NO:1.

### Claim 30 (depends on 28)

The expression vector of claim 28, further comprising an expression cassette that comprises SEQ ID NO:7.

### Claim 31

An expression vector for expression and secretion of heterologous proteins in cultured insect cells, wherein the expression vector comprises the expression cassette shown in SEQ ID NO:7.

### Claim 32 (depends on 28)

The use of the expression vector in claim 28, wherein the expression vectors are used in Drosophila cells.

### Claim 33 (depends on 28)

The use of the expression vector in claim 28, wherein the expression vectors are used in Drosophila melanogaster Schneider 2 (S2) cells.

### Claim 34

A vaccine comprising an effective amount of purified Zika virus envelope protein (E), wherein the protein is secretable into growth medium when expressed recombinantly in a host cell; and an effective amount of adjuvant, wherein the vaccine induces the production of neutralizing antibodies in human subjects.

### Claim 35 (depends on 34)

The vaccine of claim 34 wherein the E protein is recombinantly produced and expressed in insect host cells.

### Claim 36 (depends on 34)

The vaccine of claim 34 wherein the E protein is recombinantly produced and expressed in Drosophila melanogaster S2 host cells.

### Claim 37 (depends on 34)

The vaccine of claim 34, wherein the adjuvant is an aluminum-based adjuvant (Alum), a saponin-based adjuvant, monophospholipid-based adjuvant, or combination thereof.

### Claim 38 (depends on 34)

The vaccine of claim 34, for use in immunodeficient populations.

### Claim 39 (depends on 34)

The vaccine of claim 34, wherein the E protein is encoded by a nucleic acid sequence beginning at nucleotide 505 of SEQ ID NO:5.

### Claim 40 (depends on 34)

The vaccine of claim 34, wherein the E protein begins at amino acid residue 169 of SEQ ID NO:6.

### Claim 41 (depends on 34)

A method for raising a protective immune response in a person, comprising administering in a therapeutically acceptable manner a therapeutically effective amount of the vaccine of claim 34.

### Claim 42 (depends on 34)

A method of providing immune protection in humans against Zika virus induced disease comprising administering an effective amount of the vaccine of claim 34 thereby providing protection from Zika disease.