# US 10,864,232 B2 · Modified natural killer T cells, pharmaceutical compositions and uses thereof

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- Number: US 10,864,232 B2
- Document ID: US10864232B2
- Status: Granted
- Kind: B2
- Application number: 15/091,975
- Filed: 2016-04-06
- Published: 2016-10-13
- Granted: 2020-12-15
- Earliest claimed: 2015-04-07
- Assignees: Fullhope Biomedical Co., Ltd.
- Applicants: Fullhope Biomedical Co., Ltd.
- Inventors: Jan-Mou Lee; Chih-Hao Fang; Ya-Fang Cheng; Da-Tsen Wei; Kai-Yuan Jhou
- Web page: https://patentgrep.com/patent/US10864232B2

## CPC

- C12N 5/0646: {Natural killers cells [NK], NKT cells}
- A61K 40/15: Natural-killer [NK] cells; Natural-killer T [NKT] cells
- A61K 40/22: Immunosuppressive or immunotolerising
- A61K 40/416: {Antigens related to auto-immune diseases; Preparations to induce self-tolerance}
- A61P 37/00: Drugs for immunological or allergic disorders
- C12N 5/0637: {Immunosuppressive T lymphocytes, e.g. regulatory T cells or Treg}
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

The present invention provides modified natural killer T (NKT) cells, pharmaceutical compositions comprising the modified NKT cells and at least one pharmaceutically acceptable carrier or excipient, and uses of the modified NKT cells. Also disclosed herein are methods for enriching NKT cells and generating the modified natural killer T cells.

## Claims

### Claim 1

A population of natural killer T (NKT) cells carrying a phenotype of CD3+CD56+CD8+CD1d tetramer−TCR Vα24_Jα18−PD-1+CD25+Foxp3+CTLA-4+LAP+GITR−and PI-16−.

### Claim 2 (depends on 1)

The population of natural killer T cells of claim 1, wherein the NKT cells further comprise the IL-Rα+phenotype.

### Claim 3

A composition, comprising
(a) a population of natural killer T cells carrying a phenotype of CD3+CD56+CD8+CD1d tetramer−TCR Vα24_Jα18−PD-1+CD25+Foxp3+CTLA-4+LAP+GITR−and PI-16−; and
(b) a pharmaceutically acceptable carrier or excipient.

### Claim 4 (depends on 3)

The composition of claim 3, wherein the NKT cells further comprise the IL-15Rα+phenotype.

### Claim 5 (depends on 3)

The composition of claim 3, wherein the NKT cells further comprise the IL-15Rα−phenotype.

### Claim 6 (depends on 1)

A method of generating the population of natural killer T cells of claim 1, comprising the steps of
(a) culturing an enriched CD14−CD19−CD25−mononuclear cell fraction or an enriched CD4−CD14−CD16−CD19−CD25−mononuclear cell fraction with transforming growth factor beta (TGF-β), anti-CD3 antibody, and cell culture medium;
(b) contacting the cultured cell fraction from step (a) with IL-15;
(c) contacting the cultured cell fraction from step (b) with transforming growth factor beta (TGF-β);
(d) contacting the cultured cell fraction from step (c) with IL-15; and
(e) isolating the population of modified natural killer T cells carrying a phenotype of CD3+CD56+CD8+CD1d tetramer−TCR Vα24_Jα18−PD-1+CD25+Foxp3+CTLA-4+LAP+GITR−and PI-16−.

### Claim 7 (depends on 6)

The method of claim 6, wherein the cell culture medium comprises a mammalian cell culture medium and a media supplement.

### Claim 8 (depends on 7)

The method of claim 7, wherein the mammalian cell culture medium is RPMI medium 1640 and the media supplement is fetal bovine serum.

### Claim 9 (depends on 6)

The method of claim 6, wherein the cell culture medium is a hematopoietic cell medium.

### Claim 10 (depends on 6)

The method of claim 6, wherein the NKT cells further comprise the IL-Rα+phenotype.

### Claim 11 (depends on 6)

The method of claim 6, wherein the contact time of the enriched cell fraction with the transforming growth factor beta (TGF-β) or IL-15 is about 1 day to about 9 days.

### Claim 12

A population of natural killer T (NKT) cells carrying a phenotype of CD3+CD56+CD8+CD1 d-tetramer−TCR Vα24_Jα18−GITR−PI-16−CD25+Foxp3+CTLA-4+LAP+PD-1+IL-15Rα−.

### Claim 13 (depends on 6)

The method of claim 6, wherein the NKT cells further comprise the IL-15Rα- phenotype.

### Claim 14 (depends on 6)

The method of claim 6, in step (b) or (d), the concentration of IL-15 is 20 ng/ml to 160 ng/ml.