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Natural killer cell compositions and method for production of the same

A natural killer cell, a mature technology, applied in the direction of peptides, etc., can solve the problems of natural killer cells that are difficult to proliferate and are in the research and development stage

Inactive Publication Date: 2008-09-03
IND FOUND OF CHONNAM NAT UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, natural killer cells are difficult to proliferate. So far, various methods have been tried to solve this problem, but the use of interleukin-2 or interleukin-15 can only proliferate dozens of times at most, but later studies have shown that if Mixing other people's cancer cells can multiply hundreds of times
The latest proliferation method published in 2005 showed that if human cancer cells mixed with two genetic genes are co-cultured, they can multiply by 1000 times, but this method is still in the research and development stage
However, there are still safety and ethical issues in co-culturing with other people's cancer cells or using other people's genetic genes

Method used

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  • Natural killer cell compositions and method for production of the same
  • Natural killer cell compositions and method for production of the same
  • Natural killer cell compositions and method for production of the same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0106] Example 1. Mining specific genes expressed in each stage of natural killer cell differentiation in mice

[0107] (A) Method for isolating hematopoietic blasts from mouse bone marrow and method for differentiating natural killer cells

[0108] After the bone marrow cells were isolated from the whole bones of 8-12-week-old mice (C57BL / 6, purchased from Korea Reasearch Bioscience and Biotechnology, and managed in a sterile state according to animal management guidelines), they were washed with cell lysis buffer ( 0.2% NaCl, 1.6% NaCl) to remove erythrocytes, and after reacting the cells with the 2.4G2 supernatant on ice for 10 minutes, they were washed with phosphate buffered saline (buffer A) containing 2 mM EDTA. Then the cells were suspended in buffer A (1X108 / 500μl), and added with vitamin H (biotin) antibody mixture (antibodycocktail) (Mac-1, Gr-1, B220, NK 1.1, CD2, TER-119, Pharmingen ), reacted at a temperature of 4°C for 10 minutes, and then washed the cells with...

Embodiment 2

[0120] Example 2. Effect of Axl polyclonal antibody on differentiation of natural killer cells

[0121] (A) Expression of specific receptors of mature natural killer cells stimulated by Axl polyclonal antibody in co-culture with stromal cells:

[0122] During the differentiation of mouse hematopoietic blasts into natural killer cells, the precursor natural killer cells (7 days) were treated with 1 μg of Axl polyclonal antibody (Santa Cruz Biotechnology, Inc.sc-1096), and then utilized as described in Example 1 In the method of adding interleukin-15 (40ng / ml), the precursor natural killer cells and stromal cells were co-cultured for 6 days, and after the precursor natural killer cells were differentiated into mature natural killer cells, The resulting mature natural killer cells were stained with NK1.1 antibody and antibodies to natural killer cell-specific receptors (Ly49G2, Ly49A, Ly49C / F / I, Pharmingen), and then performed FACS analysis by the method described in Example 1 (B...

Embodiment 3

[0129] Example 3. Effect of Axl polyclonal antibody on the production of interferon-γ by natural killer cells

[0130] The precursor natural killer cells obtained in Example 1 (B) were co-cultured with stromal cells under the condition of Axl polyclonal antibody (500ng / ml) and interleukin-15 (10ng / ml). After the mature natural killer cells were obtained, human interleukin-2 (10 ng / ml, R&D) was used to treat and activate the activity of the mature natural killer cells. After 24 hours, use the interferon-γ enzyme-linked immunosorbent assay (ELISA) kit (BD Pharmingen) to detect the production of interferon-γ. It was found that the interferon-γ in the experimental group treated with the Axl polyclonal antibody produced more than the control group ( Figure 11 ).

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Abstract

The present invention relates to a ligand of AxI receptor tyrosine kinase used to induce the differentiation from precursor natural killer cell to mature natural killer-cell. In addition, it relates to a process for producing mature natural killer cell comprising treating hematopoietic stem cell with interleukin-7, stem cell factor and Flt3L to differentiate into precursor natural killer cell, and treating the resulting precursor natural killer cell with ligand of AxI receptor tyrosine kinase to produce mature natural killer cell.

Description

technical field [0001] The invention relates to a ligand of Axl receptor tyrosine kinase (kinase) that induces precursor natural killer cells (Natural killer cell, NK) to differentiate into mature natural killer cells. The present invention also relates to a preparation method of mature natural killer cells, which uses interleukin-7 (interleukin-7), stem cell factor (SCF) and Flt3 ligand to treat hematopoietic blasts to differentiate into precursor natural killer cells After cells, the precursor natural killer cells are treated with the ligand of Axl receptor tyrosine kinase to differentiate them into mature natural killer cells. Background technique [0002] The present invention relates to an Axl receptor tyrosine kinase (Axl Receptor tyrosine kinase, hereinafter referred to as "Axl") that induces precursor natural killer cells (pNK) to differentiate into mature natural killer cells (mNK) ) ligands. Particularly relates to a kind of preparation method of mature natural k...

Claims

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Application Information

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IPC IPC(8): C07K2/00
Inventor 姜馨植金相泫金银美李恩姬尹精高昌甫许荣真沈佑永许成范尹智然
Owner IND FOUND OF CHONNAM NAT UNIV
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