# US 2019/0376016 A1 · Personalized cellular biomanufacturing with a closed, miniature cell culture system

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- Number: US 2019/0376016 A1
- Document ID: US20190376016A1
- Status: Published application
- Kind: A1
- Application number: 16/462,753
- Filed: 2017-11-22
- Published: 2019-12-12
- Earliest claimed: 2016-11-22
- Applicants: NUtech Ventures
- Inventors: Yuguo Lei
- Web page: https://patentgrep.com/patent/US20190376016A1

## CPC

- C12M 25/14: {Scaffolds; Matrices (in general C12N5/0068)}
- C12M 29/00: {Means for introduction, extraction or recirculation of materials, e.g. pumps (pumps per se F04B)}
- C12M 33/00: {Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus (chemical or physical laboratory apparatus in general B01L, devices for taking cell samples A61B10/0045, withdrawing or distributing predetermined quantities of fluid B01L99/00)}
- C12N 5/0606: {Pluripotent embryonic cells, e.g. embryonic stem cells [ES] (embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696)}
- C12N 5/0696: {Artificially induced pluripotent stem cells, e.g. iPS}
- G01N 33/50: Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing (measuring or testing processes involving enzymes or microorganisms, compositions or test papers therefor; processes for forming such compositions, condition responsive control in microbiological or enzymological processes C12Q)
- C12N 2513/00: 3D culture
- C12N 2533/30: Synthetic polymers
- 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}
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

Closed, miniature devices and methods of using the devices for culturing cells are disclosed. Particularly, a device, and methods of using the device for manufacturing, expanding, differentiating and/or reprogramming cells for personalized medicine, such to allow for conducting medical procedures at the point-of-care, are provided.

## Claims

### Claim 1

A device for culturing cells for personalized medicine, the device comprising:
a closed housing comprising a three-dimensional (3D) hydrogel scaffold;
an inlet for introducing a cell culture medium into the housing; and
an outlet for exhausting cell culture medium from the housing.

### Claim 2 (depends on 1)

The device as set forth at claim 1 wherein the closed housing is a closed cell culture tube.

### Claim 3 (depends on 1)

The device as set forth in claim 1 wherein the closed housing has a capacity of less than 10 L.

### Claim 4 (depends on 1)

The device as set forth in claim 1 wherein the 3D hydrogel scaffold comprises at least one of poly(ethylene glycol) (PEG)-(N-isopropylacrylamide) polymers and alginate polymers.

### Claim 5

(canceled)

### Claim 6 (depends on 1)

The device as set forth in claim 1 wherein the 3D hydrogel scaffold is in a form selected from the group consisting of a sheet, fiber, hollow fiber, sphere, and combinations thereof.

### Claim 7 (depends on 1)

The device as set forth in claim 1 wherein the cells are selected from the group consisting of human primary cells, induced pluripotent stem cells (iPSCs), embryonic stem cells, and derivatives thereof, primary tumor cells, and combinations thereof.

### Claim 8

(canceled)

### Claim 9 (depends on 1)

A method of expanding cells using the device as set forth in claim 1, the method comprising:
suspending a cell solution including cells in the 3D hydrogel scaffold of the closed housing;
introducing a cell culture medium into the closed housing from the inlet; and
culturing the cells.

### Claim 10 (depends on 9)

The method as set forth in claim 9 wherein suspending the cell solution including cells comprises encapsulating cells from the cell solution into the 3D hydrogel scaffold.

### Claim 11 (depends on 9)

The method as set forth in claim 9 further comprising releasing the cultured cells from the 3D hydrogel scaffold comprising dissolving the 3D hydrogel scaffold, wherein the 3D hydrogel scaffold is dissolved by at least one of: using a chemical dissolvent selected from the group consisting of ethylenediaminetetraacetic acid (EDTA), ethylene glycol tetraacetic acid (EGTA), and an alginate lyase solution and using mechanical force.

### Claim 12

(canceled)

### Claim 13

(canceled)

### Claim 14 (depends on 9)

The method as set forth in claim 9 further comprising purifying the cultured cells by contacting the cultured cells with antibody-coated magnetic beads within the closed housing.

### Claim 15 (depends on 9)

The method as set forth in claim 9 further comprising concentrating the cultured cells by centrifuging the cultured cells or contacting the cultured cells with antibody-coated magnetic beads within the closed housing.

### Claim 16

(canceled)

### Claim 17 (depends on 9)

The device as set forth in claim 9 wherein the 3D hydrogel scaffold comprises at least one of poly(ethylene glycol) (PEG)-(N-isopropylacrylamide) polymers and alginate polymers.

### Claim 18

(canceled)

### Claim 19

(cancelled)

### Claim 20 (depends on 1)

A method of differentiating cells using the device as set forth in claim 1, the method comprising:
suspending a cell solution including cells in the 3D hydrogel scaffold of the closed housing;
introducing a cell differentiation medium into the closed housing from the inlet; and
culturing the cells.

### Claim 21 (depends on 20)

The method as set forth in claim 20 wherein suspending the cell solution including cells comprises encapsulating cells from the cell solution into the 3D hydrogel scaffold.

### Claim 22 (depends on 20)

The method as set forth in claim 20 further comprising releasing the cultured cells from the 3D hydrogel scaffold comprising dissolving the 3D hydrogel scaffold, wherein the 3D hydrogel scaffold is dissolved by at least one of: using a chemical dissolvent selected from the group consisting of ethylenediaminetetraacetic acid (EDTA), ethylene glycol tetraacetic acid (EGTA), and an alginate lyase solution and using mechanical force.

### Claim 23

(canceled)

### Claim 24

(canceled)

### Claim 25 (depends on 20)

The method as set forth in claim 20 further comprising purifying the cultured cells by contacting the cultured cells with antibody-coated magnetic beads within the closed housing.

### Claim 26 (depends on 20)

The method as set forth in claim 20 further comprising concentrating the cultured cells by centrifuging the cultured cells or contacting the cultured cells with antibody-coated magnetic beads within the closed housing.

### Claim 27

(canceled)

### Claim 28 (depends on 20)

The device as set forth in claim 20 wherein the 3D hydrogel scaffold comprises at least one of poly(ethylene glycol) (PEG)-(N-isopropylacrylamide) polymers and alginate polymers.

### Claim 29

(canceled)

### Claim 30

(canceled)

### Claim 31 (depends on 1)

A method of reprogramming cells using the device as set forth in claim 1, the method comprising:
suspending a cell solution including adult cells in the 3D hydrogel scaffold of the closed housing;
introducing a cell culture medium into the closed housing from the inlet; and
reprogramming the cells.

### Claim 32 (depends on 31)

The method as set forth in claim 31 wherein suspending the cell solution including cells comprises encapsulating cells from the cell solution into the 3D hydrogel scaffold.

### Claim 33 (depends on 31)

The method as set forth in claim 31 further comprising releasing the cultured cells from the 3D hydrogel scaffold comprising dissolving the 3D hydrogel scaffold, wherein the 3D hydrogel scaffold is dissolved by at least one of: using a chemical dissolvent selected from the group consisting of ethylenediaminetetraacetic acid (EDTA), ethylene glycol tetraacetic acid (EGTA), and an alginate lyase solution and using mechanical force.

### Claim 34

(canceled)

### Claim 35

(cancelled)

### Claim 36 (depends on 31)

The method as set forth in claim 31 further comprising purifying the cultured cells by contacting the cultured cells with antibody-coated magnetic beads within the closed housing.

### Claim 37 (depends on 31)

The method as set forth in claim 31 further comprising concentrating the cultured cells by centrifuging the cultured cells or contacting the cultured cells with antibody-coated magnetic beads within the closed housing.

### Claim 38

(canceled)

### Claim 39 (depends on 31)

The device as set forth in claim 31 wherein the 3D hydrogel scaffold comprises at least one of poly(ethylene glycol) (PEG)-(N-isopropylacrylamide) polymers and alginate polymers.

### Claim 40

(canceled)

### Claim 41

(canceled)