# US 11,110,126 B2 · Method of expanding NK cell and composition for culturing

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- Number: US 11,110,126 B2
- Document ID: US11110126B2
- Status: Granted
- Kind: B2
- Application number: 16/281,372
- Filed: 2019-02-21
- Published: 2019-06-20
- Granted: 2021-09-07
- Earliest claimed: 2015-07-10
- Assignees: Korea University Research and Business Foundation
- Applicants: Korea University Research and Business Foundation
- Inventors: Kyung-Mi Lee; Jeongwon Sohn; Seon Ah Lim; Jang-Mi Lim
- Web page: https://patentgrep.com/patent/US11110126B2

## CPC

- C12N 5/0646: {Natural killers cells [NK], NKT cells}
- A61K 35/17: Lymphocytes; B-cells; T-cells; Natural killer cells; Interferon-activated or cytokine-activated lymphocytes (when activated by a specific antigen A61K39/00)
- A61K 2039/5158: {Antigen-pulsed cells, e.g. T-cells}
- A61K 2039/585: {wherein the target is cancer}
- C12N 2501/998: Proteins not provided for elsewhere
- C12N 2501/999: Small molecules not provided for elsewhere
- 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)

## Abstract

Provided are a method of ex-vivo culture of natural killer (NK) cells by treating the cells with a reactive oxygen species (ROS) inhibitor and/or a p53 protein inhibitor; and a composition comprising the cultured NK cells. By reducing the activity of ROS and p53 proteins during ex-vivo culture, NK cells may have achieved greater expansion efficiency without altering their anti-tumor cytotoxicity.

## Claims

### Claim 1

A method of enhancing NK cell expansion, comprising:
treating natural killer (NK) cells with a p53 inhibitor; or a reactive oxygen species (ROS) inhibitor and a p53 inhibitor, wherein the p53 inhibitor is one or more selected from the group consisting of 2-(2-imino-4,5,6,7-tetrahydrobenzothiazol-3-yl)-1-p-tolylethanone hydrobromide (pifithrin-α) and pifithrin-μ.

### Claim 2 (depends on 1)

The method of claim 1, wherein NK cells are derived from human peripheral blood mononuclear cells (PBMCs).

### Claim 3 (depends on 1)

The method of claim 1, wherein the ROS inhibitor is one or more selected from the group consisting of 1-(4-hydroxy-3-methoxyphenyl)-ethanone (apocynin), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (trolox), pyrrolidine dithiocarbamate (PDTC), glutathione (GSH), catalase, manganese superoxide dismutase (MnSOD), vitamin E, and Quercetin.

### Claim 4 (depends on 1)

The method of claim 1, wherein the inhibitors include trolox and pifithrin-α in a molar ratio of 50 to 150:1.

### Claim 5 (depends on 1)

The method of claim 1, wherein the treatment includes culturing NK cells expanded from PBMCs for 1 to 3 weeks in the presence of the p53 inhibitor; or the ROS inhibitor and the p53 inhibitor.

### Claim 6 (depends on 1)

The method of claim 1, wherein the method comprises: inducing and expanding NK cells from PBMCs; and culturing the expanded NK cells for 1 to 3 weeks in the presence of the p53 inhibitor; or the ROS inhibitor and the p53 inhibitor.