# US 10,246,677 B2 · Devices, systems and methods for the production of humanized gut commensal microbiota

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- Number: US 10,246,677 B2
- Document ID: US10246677B2
- Status: Granted
- Kind: B2
- Application number: 15/893,028
- Filed: 2018-02-09
- Published: 2018-06-21
- Granted: 2019-04-02
- Earliest claimed: 2016-07-28
- Applicants: Bobban Subhadra
- Inventors: Bobban Subhadra
- Web page: https://patentgrep.com/patent/US10246677B2

## 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 system for the ex vivo production of human gut microbiota, comprising:
a tortuous, tubular ex vivo culture network comprising-hollow cartridges for culturing human gut microbiota, wherein the hollow cartridges are in the fluid communication and removably attached to the tortuous tubular ex vivo culture network and comprise human gut microbiota and metabolites;
a humidifying system in fluid communication with the tortuous tubular ex vivo culture network to maintain physiological humidity inside the tortuous tubular ex vivo culture network;
a temperature regulation system in contact with the tortuous tubular ex vivo culture network to regulate the temperature inside the tortuous tubular ex vivo culture network to physiological temperature; and
a hollow functional screening portal comprising extracellular matrix proteins, human cells, and metabolites generated by the microbiota in the tortuous tubular ex vivo culture network, wherein the screening portal receives oxygen and other gases for culturing the human cells from a gas system in fluid communication with the tortuous tubular ex vivo culture network, wherein the functional screening portal is separated from the microbiota column with a semi-permeable membrane; and
a tunable, gas gradient of 0.8-2.0% oxygen or 2-60 mm Hg in the tortuous tubular ex vivo culture network.

### Claim 2 (depends on 1)

The system of claim 1, wherein the gas system comprises an oxygen sensor in fluid communication with the tortuous, tubular ex vivo culture network.

### Claim 3 (depends on 1)

The system of claim 1, further comprising a nutrient medium for culturing human gut microbiota in the tortuous tubular culture network.

### Claim 4 (depends on 1)

The system of claim 1, wherein the hollow cartridges have an interior surface coated with extracellular matrix proteins.

### Claim 5 (depends on 4)

The system of claim 4, wherein the extracellular proteins are covered with a layer of cells of the intestine.

### Claim 6 (depends on 4)

The system of claim 4, wherein the extracellular matrix proteins comprise one or more proteins selected from the group consisting of collagens, elastins, fibronectins, laminins and combinations thereof.

### Claim 7 (depends on 1)

The system of claim 1, wherein the cartridge further comprises a substrate.

### Claim 8 (depends on 7)

The system of claim 7, wherein the substrate is selected from the group consisting of glass wool, rockwool, porous sponge, perforated sponge, or combinations thereof.

### Claim 9 (depends on 3)

The system of claim 3, wherein the nutrient medium comprises: organic nitrogen sources, protein sources, carbohydrate sources, metal salts, intestinal secretions, enzymes, vitamins, and trace minerals maintained at a slightly neutral to acidic pH.

### Claim 10 (depends on 1)

The system of claim 1, wherein the functional screening portal comprises a cell-growing surface with a cell culture media, and a semi-permeable membrane that allows diffusion of only metabolites from the plurality of cartridges.

### Claim 11 (depends on 1)

The system of claim 1, wherein an interior environment of the tortuous tubular culture network mimics the interior environment of human intestine.

### Claim 12 (depends on 1)

The system of claim 1, wherein the hollow cartridge comprises a sample of gut microbiota suitable for transplant into a subject in need thereof.

### Claim 13 (depends on 1)

The system of claim 1, wherein the hollow cartridge comprises a sample of probiotic bacteria suitable for transplant into a subject in need thereof.