# US 2013/0156809 A1 · Recombinant Bacterium and Uses Thereof

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- Number: US 2013/0156809 A1
- Document ID: US20130156809A1
- Status: Published application
- Kind: A1
- Application number: 13/811,690
- Filed: 2011-07-28
- Published: 2013-06-20
- Earliest claimed: 2010-07-28
- Assignees: National Research Council of Canada
- Inventors: Subash Sad; Lakshmi Krishnan; Valeria Alcon; Kevin G. Young; Fanny Tzelepis
- Web page: https://patentgrep.com/patent/US20130156809A1

## CPC

- C07K 14/77: Ovalbumin
- A61K 39/0005: {Vertebrate antigens (from snakes A61K39/38)}
- A61K 39/0011: {Cancer antigens}
- A61K 39/001106: {Her-2/neu/ErbB2, Her-3/ErbB3 or Her 4/ErbB4}
- A61K 39/001156: {Tyrosinase and tyrosinase related proteinases [TRP-1 or TRP-2]}
- A61K 39/001186: {MAGE}
- A61K 39/001191: {Melan-A/MART}
- A61K 39/001192: {Glycoprotein 100 [Gp100]}
- A61K 39/12: Viral antigens
- A61K 40/11: T-cells, e.g. tumour infiltrating lymphocytes [TIL] or regulatory T [Treg] cells; Lymphokine-activated killer [LAK] cells
- A61K 40/42: Cancer antigens
- A61K 40/4562: {Salmonella; Shigella}
- A61K 40/46: Viral antigens
- A61P 31/04: Antibacterial agents
- A61P 31/12: Antivirals
- A61P 37/04: Immunostimulants
- C07K 14/195: from bacteria
- C07K 14/4713: {Autoimmune diseases, e.g. Insulin-dependent diabetes mellitus, multiple sclerosis, rheumathoid arthritis, systemic lupus erythematosus; Autoantigens}
- C07K 14/4748: {Tumour specific antigens; Tumour rejection antigen precursors [TRAP], e.g. MAGE}
- C07K 14/70596: {Molecules with a "CD"-designation not provided for elsewhere}
- C07K 14/71: for growth factors; for growth regulators
- C12N 1/20: Bacteria; Culture media therefor
- C12N 9/0059: {Catechol oxidase (1.10.3.1), i.e. tyrosinase}
- C12N 9/2462: {Lysozyme (3.2.1.17)}
- C12N 9/6494: {Neprilysin (3.4.24.11), i.e. enkephalinase or neutral-endopeptidase 24.11}
- C12Y 110/03001: Catechol oxidase (1.10.3.1), i.e. tyrosinase
- C12Y 304/24011: Neprilysin (3.4.24.11), i.e. enkephalinase or neutral endopeptidase 24.11
- A61K 2039/522: {avirulent or attenuated}
- A61K 2039/523: {expressing foreign proteins}
- A61K 2239/31: characterized by the route of administration
- A61K 2239/38: characterised by the dose, timing or administration schedule
- C07K 2319/036: targeting to the medium outside of the cell, e.g. type III secretion
- C07K 2319/35: containing a fusion for enhanced stability/folding during expression, e.g. fusions with chaperones or thioredoxin
- C12N 2760/10034: 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)
- C12Y: ENZYMES
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

The present invention relates to a recombinant bacterium expressing an antigen that is translocated to the cytosol of a host organism, and uses thereof. To this end, the present invention provides a recombinant bacterium comprising a nucleic acid encoding an antigen that is translocated to the cytosol of a host cell utilizing Type III secretion system. The recombinant bacterium is generally chosen from intracellular pathogens that reside in the phagosome and fail to induce rapid T cell activation. The translocated antigen may be a viral antigen, a bacterial antigen, or a tumour antigen. Methods of imparting immunity using the recombinant bacterium are also provided.

## Claims

### Claim 1

A recombinant bacterium, comprising a nucleic acid encoding an antigen that is translocated to the cytosol of a host cell.

### Claim 2 (depends on 1)

The recombinant bacterium of claim 1, wherein the bacterium is an intracellular bacterium such as Salmonella, Mycobacteria, Brucella, or Leishmania.

### Claim 3 (depends on 1)

The recombinant bacterium of claim 1, wherein the bacterium is Salmonella.

### Claim 4 (depends on 1)

The recombinant bacterium of claim 1, wherein the antigen is a viral antigen, a bacterial antigen, or a tumour antigen.

### Claim 5 (depends on 1)

The recombinant bacterium of claim 1, wherein the antigen is a fusion protein comprising an antigen and a translocation domain from a type III secretion system.

### Claim 6 (depends on 5)

The recombinant bacterium of claim 5, wherein the translocation protein is YopE, SopE, SptP, or a fragment thereof.

### Claim 7 (depends on 5)

The recombinant bacterium of claim 5, wherein the fusion protein further comprises a chaperone.

### Claim 8 (depends on 7)

The recombinant bacterium of claim 7, wherein the chaperone is derived from type Ill secretion systems.

### Claim 9 (depends on 8)

The recombinant bacterium of claims 8, wherein the chaperone is SycE or HSP70.

### Claim 10 (depends on 1)

The recombinant bacterium of claim 1, wherein the nucleic acid is comprised in a vector.

### Claim 11 (depends on 10)

The recombinant bacterium of claim 10, wherein the vector is a pHR vector.

### Claim 12 (depends on 10)

The recombinant bacterium of claim 10, wherein the vector is pHR-241.

### Claim 13 (depends on 1)

The recombinant bacterium of claim 1, wherein the antigen is tyrosinase related protein 2 (TRP-2), MART-1, melanoma associated antigen 1 (MAGE1), or Her-2/neu, gp100, or other viral or bacterial antigens.

### Claim 14 (depends on 1)

A method of imparting immunity against naturally-occurring bacterium or virus in a subject, the method comprising administering the recombinant bacterium of claim 1 to said subject.

### Claim 15 (depends on 13)

A method of imparting immunity against tumors in a subject, the method comprising administering the recombinant bacterium of claim 13 to said subject.

### Claim 16 (depends on 14)

The method of claim 14, wherein the recombinant bacterium is administered through intravenous, oral or subcutaneous routes of immunization.

### Claim 17 (depends on 1)

The use of the recombinant bacterium of claim 1 as a vaccine.

### Claim 18 (depends on 15)

The method of claim 15, wherein the recombinant bacterium is administered through intravenous, oral or subcutaneous routes of immunization.