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A kind of trophoblast cell and its application in expanding human NK cells

A technology of trophoblast cells and NK cells, applied in trophoblast cells and its application in the expansion of human NK cells, to achieve the effects of reducing potential safety hazards, good safety, and good anti-tumor activity

Active Publication Date: 2022-06-24
CELARTICS BIOPHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, judging from the killing data of NK cells and the clinical performance of NK cells, there is still a lot of room for improvement in the activity of NK cells

Method used

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  • A kind of trophoblast cell and its application in expanding human NK cells
  • A kind of trophoblast cell and its application in expanding human NK cells
  • A kind of trophoblast cell and its application in expanding human NK cells

Examples

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preparation example Construction

[0053] The present application provides a method for preparing the above-mentioned trophoblast cells, the preparation method specifically includes the following steps: constructing the above-mentioned cytokine lentiviral vector, co-introducing the above-mentioned cytokine lentivirus vector with an auxiliary plasmid into mammalian cells to obtain the above-mentioned cytokine lentivirus; The factor lentivirus infects the host cells and transduces them to obtain trophoblast cells.

[0054] In addition, the present application also provides an NK cell expansion medium, the medium comprising the above-mentioned trophoblast cells. Further, the NK cell culture medium also includes any one or a combination of at least two of basal medium, serum, plasma or antibiotics.

[0055] In addition, the present application also provides the preparation method of the above-mentioned trophoblast cells, cytokine lentiviral vectors, cytokine lentiviruses, trophoblast cells or the application of the...

preparation example 1-8

[0060] Preparation Examples 1-8 respectively provide a method for constructing a cytokine lentiviral vector. The difference lies in the different types of cytokines in the prepared cytokine lentiviral vector. The specific operation steps are as follows:

[0061] (1) Obtain the coding sequence information of cytokines, design primers and conduct primer synthesis by a third-party company;

[0062] (2) Using the cDNA of lymph node cells as a template, clone the coding region of cytokines, and insert the lentiviral vector EcoRI-XbaI or XhoI-XbaI site after enzyme digestion, and transform into E.coli; after correct sequencing, shake the bacteria , extraction to obtain cytokine lentiviral vector;

[0063] Among them, the way to obtain the information of cytokine coding sequence is the NCBI database; lymph node cells are commercially available.

[0064] The cytokine species corresponding to the above Preparation Examples 1-8, their coding sequence information, and their correspond...

preparation example 9-16

[0068] Preparation Examples 9-16 respectively provide a production method of a cytokine lentiviral vector. The difference lies in the different types of cytokine lentiviral vectors produced. The specific operation steps are as follows:

[0069] (1) Co-transfect 293 cells (human renal epithelial cell line) with the cytokine lentiviral vectors obtained in Preparation Examples 1-8 and the mixed packaging plasmids through the transfection reagent-calcium phosphate transfection reagent, respectively, to obtain the co-transfected 293 cells; the co-transfected 293 cells were incubated at 37°C in CO 2 Culture in an incubator, take the cultured supernatant at 24h, 48h, and 72h, respectively, to obtain the supernatant of the co-transfected 293 cells. Among them, the mixed packaging plasmids include the expression plasmids of VSV-G, Gag-Pol and Rev.

[0070] (2) Collect the supernatant of the co-transfected 293 cells cultured in step (1), and centrifuge the collected supernatant at 4°C...

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Abstract

The application relates to the technical field of genetic engineering, and specifically discloses a trophoblast cell and its application in expanding human NK cells. The trophoblast cells express cytokines, and the cytokines include CD80, CD70, CD86, 4‑1BBL, CD40, CD58, CD83 and MICA; the trophoblast cells are NIH 3T3 cells; and are used to prepare the trophoblast cells The cytokine lentiviral vector and the cytokine lentivirus prepared by using the cytokine lentiviral vector; and the NK cell expansion medium prepared by using the trophoblast cells; and the application of the trophoblast in expanding NK cells. The NK cells prepared in this application have better safety and ideal killing data.

Description

technical field [0001] The present application relates to the technical field of genetic engineering, more specifically, it relates to a trophoblast cell and its application in expanding human NK cells. Background technique [0002] Natural killer cells (NK cells) are lymphocytes produced from CD34+ hematopoietic progenitor cells in the bone marrow. NK cells are mainly found in the blood, liver, spleen, and bone marrow, but factors such as inflammation, infection, etc., can trigger NK cells to migrate into almost any tissue. NK cells are an important type of immune cells in the human immune system. They can quickly respond to and kill cells infected with foreign pathogens and cells with malignant transformation caused by stress, and play a key role in the body's anti-virus and anti-tumor processes. [0003] In the related art, NK cells mainly mediate cytotoxicity through the following mechanisms. Of these, the most studied pathway is degranulation, in which NK cells releas...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/10C12N15/867C12N7/01C12N5/0783C07K14/705C07K14/74
CPCC12N5/0656C12N15/86C12N7/00C12N5/0646C07K14/70532C07K14/70575C07K14/70528C07K14/70578C07K14/70503C07K14/70539C12N2740/15021C12N2740/15043C12N2502/13
Inventor 张欢于洋程铧
Owner CELARTICS BIOPHARMA CO LTD
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