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Nucleic acid coded CAR-T cells, and preparation method and application thereof

A cell and coding technology, applied in the field of medical diagnosis and treatment, can solve problems such as strong toxic and side effects

Active Publication Date: 2020-07-07
苏州朴衡科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At the same time, this method can effectively solve the complicated problem of CAR-T cell preparation. For the mutation of surface characteristic molecules during the development of cancer cells, the type of modified antibody can be adjusted in time to respond, effectively improve the target rate, and overcome the serious off-target of CAR-T cell therapy. , strong toxic and side effects

Method used

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  • Nucleic acid coded CAR-T cells, and preparation method and application thereof
  • Nucleic acid coded CAR-T cells, and preparation method and application thereof
  • Nucleic acid coded CAR-T cells, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] (1) will 1×10 5 T cells were mixed with cholesterol-modified DNA in the medium to make the final concentration of DNA reach 10 μmol / L, at 37°C, 5% CO 2 Incubate for 1 hour to obtain cells modified with single-stranded DNA;

[0064] (2) After washing the cells modified with single-stranded DNA in step (1), add the antibody modified with complementary strand DNA, mix well, and store at 37°C, 5% CO 2 Incubate for 1 hour to obtain the CAR-T cells encoded by the nucleic acid.

Embodiment 2

[0066] (1) will 1×10 5 T cells were mixed with cholesterol-modified DNA in culture medium at 37°C, 5% CO 2 Incubate for 1 hour to obtain cells modified with single-stranded DNA;

[0067] (2) After washing the cells modified with single-stranded DNA in step (1), add the antibody modified with complementary strand DNA, mix well, and store at 37°C, 5% CO 2 Incubate for 1 hour to obtain the CAR-T cells encoded by the nucleic acid.

[0068] Adjusting the amount of DNA added in step (1) can effectively control the amount of modification of cell surface molecules. Will 1×10 5 T cells were mixed with cholesterol-modified DNA in the medium so that the final concentrations of DNA reached 0, 2, 4, 8, and 16 μmol / L, respectively, at 37°C and 5% CO 2 Incubate for 1 hour to obtain cells modified with single-stranded DNA. Such as figure 2 As shown, the amount of DNA added will directly affect the modification of the cell surface; at the same time, CAR-T cells with different targeting ...

Embodiment 3

[0070] The CAR-T cells encoded by the nucleic acid prepared in Experimental Example 1 of the present invention were used to kill the tumor cell K562 as the treatment group.

[0071] In this experiment, in vitro cell experiments were used as cancer treatment models. By adding nucleic acid-encoded CAR-T cells to tumor cell K562, observing the adhesion of CAR-T cells to tumor cell K562, and evaluating the targeting effect of nucleic acid-encoded CAR-T cells.

[0072] (1) Modeling

[0073] Wash the tumor cell K562 expressing the target molecule on the surface according to 2×10 per well 4 Cells were incubated in a 96-well plate as the tumor to be treated.

[0074] (2) experiment

[0075] Will 1×10 5 CAR-T cells encoded by a nucleic acid were added to the tumor cell K562 in the treatment group, mixed evenly, and kept at 37°C, 5% CO 2 Incubation. Before adding nucleic acid-encoded CAR-T cells, the tumor cell K562 was stained with Hoechst for later observation. After incubation...

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Abstract

The invention discloses a preparation method of nucleic acid coded CAR-T cells. The preparation method of the nucleic acid coded CAR-T cells comprises the steps of: directly mixing DNA modified with cholesterol with cells, and incubating at 37 DEG C; and, adding a complementary DNA chain modified with an antibody, uniformly mixing, and incubating at 37 DEG C, so that nucleic acid coded CAR-T cellsare obtained. The preparation method is simple in reaction condition, few in synthesis step, short in time consumption and low in production cost. When being used for cancer treatment, the nucleic acid coded CAR-T cells are good in biocompatibility, little in toxic and side effect, high in controllability and flexible, and can effectively kill tumour cells. The CAR-T cells in the invention have potential application value for prophase cancer treatment.

Description

technical field [0001] The invention belongs to the field of medical diagnosis and treatment, and relates to an immunotherapy cell and a preparation method thereof, in particular to an immune cell capable of precisely targeting tumor cells and a preparation method thereof. Background technique [0002] Cancer is the most common type of malignant tumors, and its fatality rate has jumped to the first place. According to the statistics of the World Health Organization, about 4 million people die of cancer in the world every year. If the treatment is not timely, the cancer cells will continue to gnaw at the body until they take away life. Traditionally, the commonly used treatment methods are surgery, chemotherapy and radiotherapy, etc., but the results are often unsatisfactory, and the 5-year survival rate of patients receiving treatment is only about 50%. [0003] Immunotherapy aims to activate the human immune system and rely on its own immune function to kill tumor cells. ...

Claims

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Application Information

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IPC IPC(8): C12N5/10A61K35/17A61P35/00A61P35/02
CPCC12N5/0636C07K14/7051A61K35/17A61P35/00A61P35/02C12N2510/00
Inventor 裴昊姚晓玮李丽
Owner 苏州朴衡科技有限公司
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