# US 9,040,302 B2 · Genetically modified Streptococcus thermophilus bacterium

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 9,040,302 B2
- Document ID: US9040302B2
- Status: Granted
- Kind: B2
- Application number: 13/697,860
- Filed: 2011-05-19
- Published: 2013-10-03
- Granted: 2015-05-26
- Earliest claimed: 2010-05-19
- Assignees: North Carolina State University; University of Virginia Patent Foundation
- Applicants: Todd R. Klaenhammer; Richard L. Guerrant; Glynis L. Kolling; Evelyn Durmaz; Michael P. Timko; Cirle Alcantara Warren
- Inventors: Todd R. Klaenhammer; Richard L. Guerrant; Glynis L. Kolling; Evelyn Durmaz; Michael P. Timko; Cirle Alcantara Warren
- Web page: https://patentgrep.com/patent/US9040302B2

## CPC

- A61K 38/164: {from bacteria}
- A23C 9/1238: {using specific L. bulgaricus or S. thermophilus microorganisms; using entrapped or encapsulated yoghurt bacteria; Physical or chemical treatment of L. bulgaricus or S. thermophilus cultures; Fermentation only with L. bulgaricus or only with S. thermophilus}
- A23L 33/135: Bacteria or derivatives thereof, e.g. probiotics
- A61K 35/74: Bacteria (therapeutic use of a bacterial protein A61K38/00)
- A61P 1/04: for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
- A61P 31/04: Antibacterial agents
- C12N 1/06: Lysis of microorganisms
- C12P 21/02: having a known sequence of two or more amino acids, e.g. glutathione
- A61K 2035/11: {Medicinal preparations comprising living procariotic cells}
- 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)
- A23C: DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; PREPARATION THEREOF (preservation thereof A23B11/00)
- A23L: FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF (preservation thereof A23B)
- 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)
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}

## Abstract

Methods and compositions for targeted delivery of biotherapeutics are provided. The compositions comprise bile-sensitive St. thermophilus bacteria modified to release a biotherapeutic agent following bile exposure. Biotherapeutic agents released by the St. thermophilus bacteria disclosed herein include AQ and AQR rich peptides. Methods of the invention comprise administering to a subject a St. thermophilus bacterium modified to release a biotherapeutic agent following bile exposure. Administration of the St. thermophilus bacterium promotes a desired therapeutic response. The bacterium may be modified to express and release AQ or AQR rich peptides which subsequently inhibit cellular apoptosis or reduce mucosal damage. Thus, methods of the invention find use in treating or preventing a variety of gastrointestinal disorders including C. difficile infection and antibiotic-associated diarrhea.

## Claims

### Claim 1

A Streptococcus thermophilus bacterium that is genetically modified to express a heterologous polypeptide comprising an amino acid sequence having at least 90% sequence identity to the amino acid sequence as set forth in SEQ ID NO: 2 or 8, wherein said heterologous polypeptide is released from said bacterium following exposure to bile.

### Claim 2 (depends on 1)

The bacterium of claim 1, wherein said heterologous polypeptide is a therapeutic polypeptide.

### Claim 3 (depends on 1)

The bacterium of claim 1, wherein said heterologous peptide comprises
an amino acid sequence selected from the group consisting of: SEQ ID NO: 2 and 8.

### Claim 4 (depends on 1)

The bacterium of claim 1, wherein said bacterium does not survive said bile exposure.

### Claim 5 (depends on 1)

The bacterium of claim 1, wherein said Streptococcus thermophilus is selected from the group consisting of:
a) Streptococcus thermophilus NCK2071, having been deposited under ATCC Accession Number PTA-11889; and
b) a probiotic Streptococcus thermophilus bacterium.

### Claim 6

A method of making a Streptococcus thermophilus bacterium, said method comprising genetically modifying said bacterium to express a heterologous polypeptide comprising an amino acid sequence having at least 90% sequence identity to the amino acid sequence as set forth in SEQ ID NO: 2 or 8, wherein said polypeptide is released from said bacterium following exposure to bile.

### Claim 7 (depends on 6)

The method of claim 6, wherein said heterologous polypeptide comprises an amino acid sequence selected from the group consisting of:
SEQ ID NO: 2 and 8.

### Claim 8 (depends on 1)

A pharmaceutical composition comprising the bacterium according to claim 1.