# US 10,767,157 B2 · Devices, systems and methods for the production of humanized gut commensal microbiota

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- Number: US 10,767,157 B2
- Document ID: US10767157B2
- Status: Granted
- Kind: B2
- Application number: 15/598,488
- Filed: 2017-05-18
- Published: 2018-02-01
- Granted: 2020-09-08
- Earliest claimed: 2016-07-28
- Applicants: Bobban Subhadra
- Inventors: Bobban Subhadra
- Web page: https://patentgrep.com/patent/US10767157B2

## CPC

- C12N 1/20: Bacteria; Culture media therefor
- A61K 9/0031: {Rectum, anus}
- A61K 9/0053: {Mouth and digestive tract, i.e. intraoral and peroral administration}
- A61K 35/74: Bacteria (therapeutic use of a bacterial protein A61K38/00)
- C12M 23/02: {Form or structure of the vessel (large containers B65D88/00)}
- C12M 23/06: {Tubular (C12M23/08, C12M23/16 take precedence)}
- C12M 23/20: {Material Coatings (immunocoatings C12M25/00)}
- C12M 23/42: {Integrated assemblies, e.g. cassettes or cartridges}
- C12M 25/10: {Hollow fibers or tubes (hollow fiber modules in general B01D63/02)}
- C12M 41/12: {of temperature (controlling the temperature of chemical or physical processes B01J19/0013, heating or cooling apparatus for laboratory use B01L7/00)}
- C12M 41/14: {Incubators; Climatic chambers (per se B01L1/00)}
- C12M 41/34: {of gas}
- C12N 1/04: Preserving or maintaining viable microorganisms (immobilised microorganisms C12N11/00)
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- A61K 2035/11: {Medicinal preparations comprising living procariotic cells}
- C12N 1/205: Bacterial isolates
- C12N 2795/00021: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2795/00032: Use of virus as therapeutic agent, other than vaccine, e.g. as cytolytic agent
- C12N 2795/00051: Methods of production or purification of viral material
- C12R 2001/01: Bacteria or Actinomycetales {; using bacteria or Actinomycetales}
- C12R 2001/145: Clostridium
- C12R 2001/19: Escherichia coli
- C12R 2001/46: Streptococcus {; Enterococcus; Lactococcus}
- 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
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- 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)
- C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS}
- C12R: INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

One embodiment provides a commensal gut production platform for ex vivo production of human gut commensal microbiota. Another embodiment provides devices, systems and methods for ex vivo culturing of gut microflora in a system that mimics the human gut environment. The culturing of the commensal microbiota in the disclosed systems produces gut microbiota having defined characteristics and properties that can be exploited to treat various conditions in a subject.

## Claims

### Claim 1

A method of ex vivo production of human commensal gut microbiota expressing a synthetic gene, the method comprising:
screening a stool sample obtained from a healthy human subject comprising human commensal gut microbiota to detect and remove ova, parasites, and viruses;
mixing the screened human commensal gut microbiota with a synthetic ex vivo culture medium to prepare a seed culture, wherein the culture medium comprises nutrients and metabolites present in the human intestine;
infusing the seed culture into catridges coated with human cells, tissue extracts, adhesive proteins, or commensal colonizing factors;
culturing the human commensal gut microbiota in the catridges, wherein the culturing comprises maintaining the temperature and gas gradients of the culture medium so as to be similar to the human gut, wherein the gas gradient comprises dissolved oxygen;
modifying the cultured human commensal gut microbiota, wherein the modifying comprises infusing a synthetic gene into the culture medium in a concentration effective to be taken up by the cultured human commensal gut microbiota and
harvesting the cultured human commensal gut microbiota.

### Claim 2 (depends on 1)

The method of claim 1, wherein the cultured human commensal gut microbiota comprising the synthetic gene is able to secrete a therapeutic protein, peptide, or lipid.

### Claim 3 (depends on 2)

The method of claim 2, wherein the therapeutic protein or peptide is bone marrow protein (BMP), erythropoietin (EPO), granulocyte-colony-forming factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), interferon alpha, interferon beta, interferon gamma, interleukin 2 (IL-2), interleukin 11 (IL-11), mammary-associated serum amyloid protein (M-SAA), insulin, glucose-dependent insulinotropic peptide (GIP), glucagon-like peptide-1 (GLP-1), xenin, leptin, diacetylghrelin, ghrelin, or humanin.

### Claim 4 (depends on 1)

The method of claim 1, further comprising:
purifying the harvested human commensal gut microbiata and
sequencing the purified harvested human commensal gut microbiota to determine the relative proportions and diversity of microbiota therein.

### Claim 5 (depends on 1)

The method of claim 1, further comprising:
storing the harvested human commensal gut microbiota as a freeze-dried powder.

### Claim 6 (depends on 2)

The method of claim 2, further comprising purifying the harvested human commensal gut microbiota and administering the purified harvested human commensal gut microbiota into the gastrointestinal tract of a patient in need thereof.

### Claim 7 (depends on 6)

The method of claim 6, wherein the patient in need has a gut infection caused by enteropathogenic Escherichia coli (EPEC), enterotoxigenic Escherichia coli (ETEC), or enteroinvasive Escherichia coli (EIEC).

### Claim 8 (depends on 6)

The method of claim 6, wherein the patient in need has necrotizing enterocolitis.