# US 11,298,385 B2 · Microorganism for delivering drug for treatment of gastrointestinal disease, which expresses and secretes P8 protein, and pharmaceutical composition for preventing or treating gastrointestinal disease, which includes the same

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- Number: US 11,298,385 B2
- Document ID: US11298385B2
- Status: Granted
- Kind: B2
- Application number: 16/471,940
- Filed: 2018-10-16
- Published: 2019-10-31
- Granted: 2022-04-12
- Earliest claimed: 2018-01-09
- Assignees: Cell Biotech Co., Ltd.
- Applicants: Cell Biotech Co., Ltd.
- Inventors: Myung Jun Chung
- Web page: https://patentgrep.com/patent/US11298385B2

## CPC

- A61K 35/747: Lactobacilli, e.g. L. acidophilus or L. brevis
- A61K 35/744: Lactic acid bacteria, e.g. enterococci, pediococci, lactococci, streptococci or leuconostocs
- A61K 35/745: Bifidobacteria
- A61K 38/164: {from bacteria}
- C07K 14/47: from mammals
- C12N 1/20: Bacteria; Culture media therefor
- C12N 1/205: Bacterial isolates
- C12N 15/625: {containing a sequence coding for a signal sequence}
- C12N 15/74: Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
- C12N 15/746: {for lactic acid bacteria (Streptococcus; Lactococcus; Lactobacillus; Pediococcus; Enterococcus; Leuconostoc; Propionibacterium; Bifidobacterium; Sporolactobacillus)}
- A61K 35/74: Bacteria (therapeutic use of a bacterial protein A61K38/00)
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61K 2039/522: {avirulent or attenuated}
- C12R 2001/23: Lactobacillus acidophilus
- C12R 2001/24: Lactobacillus brevis
- C12R 2001/245: Lactobacillus casei
- 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)
- 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)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12R: INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS

## Abstract

The present invention relates to a microorganism for drug delivery, which has been transformed with a gene construct comprising a therapeutically active peptide, which can be delivered safely into the intestines through oral administration, and which can express and secrete the therapeutic peptide in the gastrointestinal tract, and also relates to a pharmaceutical composition for prevention or treatment of gastrointestinal disease, which includes the same. The present invention is directed to a lactic acid bacteria drug delivery system capable of overexpressing and secreting a lactic acid bacteria-derived anticancer protein, which is developed by introducing the anticancer protein into a lactic acid bacteria expression and secretion system. It is expected that the lactic acid bacteria drug delivery system will be widely used as a natural protein therapeutic agent against gastrointestinal disease in the medical field.

## Claims

### Claim 1

A Pediococcus pentosaceus strain for treating colorectal cancer, comprising a DNA construct that comprises:
an exogenous promoter;
a polynucleotide encoding a P8 protein operably linked to the exogenous promoter and having the nucleotide sequence represented by SEQ ID NO: 1; and
a gene encoding a secretion signal peptide,
wherein the Pediococcus pentosaceus strain is capable of expressing and secreting the P8 protein in a gastrointestinal tract, and
the exogenous promoter is at least one promoter selected from the group consisting of SEQ ID NO: 3 to SEQ ID NO: 8.

### Claim 2 (depends on 1)

The Pediococcus pentosaceus strain of claim 1, wherein the secretion signal peptide is a USP45 secretion signal peptide, Usp45 N4, or a Lactobacillus brevis S-layer protein signal peptide.

### Claim 3 (depends on 2)

The Pediococcus pentosaceus strain of claim 2, wherein the secretion signal peptide is the USP45 secretion signal peptide.

### Claim 4 (depends on 1)

The Pediococcus pentosaceus strain of claim 1, wherein the DNA construct further comprises
a second secretion signal peptide and a heterologous nucleic acid sequence encoding a second therapeutic peptide,
wherein the second secretion signal peptide and the heterologous nucleic acid sequence are disposed downstream of the exogenous promoter.

### Claim 5 (depends on 1)

A pharmaceutical composition for treatment of colorectal cancer, comprising the Pediococcus pentosaceus strain of claim 1, which is capable of producing and secreting a P8 protein having the amino acid sequence represented by SEQ ID NO: 2.

### Claim 6 (depends on 1)

The Pediococcus pentosaceus strain of claim 1, wherein the Pediococcus pentosaceus strain is deposited in the Korean Culture Center of Microorganisms (KCCM) under the accession number KCCM12181P.

### Claim 7 (depends on 5)

The pharmaceutical composition of claim 5, wherein the Pediococcus pentosaceus strain is deposited in the Korean Culture Center of Microorganisms (KCCM) under the accession number KCCM12181P.

### Claim 8 (depends on 4)

The Pediococcus pentosaceus strain of claim 4, wherein the exogenous promoter includes a first promoter and a second promoter, and the second promoter is identical to the first promoter.

### Claim 9 (depends on 4)

The Pediococcus pentosaceus strain of claim 4, wherein the exogenous promoter includes a first promoter and a second promoter, and the second promoter is different from the first promoter.