A method for in vitro culture of T cells loaded with PD-1 antibody, its cell preparation and its application

A culture method and PD-1 technology, applied in the field of in vitro culture of T cells loaded with PD-1 antibody, can solve problems such as the absence of PD-1 antibody drugs and the toxic and side effects of PD-1 antibody drugs, so as to enhance lethality and avoid Loss of function effects

Active Publication Date: 2020-10-30
广州筑康生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] However, the efficacy of PD-1 antibody can only be used for some patients, because the tumor produces PD-L1, and for other tumors that are not caused by PD-L1, or the PD-1 antibody drug has no effect, and PD -1 Antibody Drugs Have Great Toxic Side Effects

Method used

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  • A method for in vitro culture of T cells loaded with PD-1 antibody, its cell preparation and its application
  • A method for in vitro culture of T cells loaded with PD-1 antibody, its cell preparation and its application
  • A method for in vitro culture of T cells loaded with PD-1 antibody, its cell preparation and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] The method for obtaining killer T cells is as follows.

[0057] Take 150ml of peripheral blood, or collect umbilical cord blood, and use lymphocyte separation medium and gradient centrifugation to separate mononuclear cells (mainly including lymphocytes, as well as other white blood cells and a small amount of red blood cells).

[0058] first training

[0059] For the initial culture, the isolated mononuclear cells are mixed with anti-A medium, put into a cell culture square bottle, and placed at 37°C, 5% CO 2 cultured in a constant temperature and humidity carbon dioxide incubator.

[0060] The components of A anti-medium are as follows:

[0061] Complete medium, CD3 monoclonal antibody, interleukin-2, vitamin B6, interferon-γ, interleukin-1b, interleukin-4, L-glutamine, sodium propupuprate and plasma protein.

[0062] Wherein, the final concentration of CD3 monoclonal antibody is 100ng / ml; preferably, in the anti-A culture medium, the final concentration of interle...

Embodiment 2

[0076] The curative effect of the cell preparation provided in Example 1 for patients with cervical cancer.

[0077] The PET-CT examination results of the cervical cancer patient on December 29, 2017 were:

[0078] 1. After cervical cancer surgery, the uterus and bilateral appendages are absent; there is no sign of malignant tumor in the vaginal stump;

[0079] 2. The left supraclavicular fossa lymph node metastases seen in the last imaging (July 11, 2017) disappeared in this imaging, the left supraclavicular fossa soft tissue was disordered, and the metabolism was slightly increased, which was considered to be a change after treatment;

[0080] 3. Several slightly enlarged lymph nodes were removed from the retroperitoneum in the middle and upper abdomen, and the metabolism was slightly increased. It was considered lymph node inflammation, and the metabolism decreased compared with the previous imaging;

[0081] 4. A nodular hypermetabolism lesion in the upper part of the lef...

Embodiment 3

[0097] The curative effect of the cell preparation provided in Example 1 on leukemia patients was verified.

[0098] The leukemia patient was unwell at the end of 2017. He went to the local hospital for examination and was diagnosed with monocytic leukemia (M5). After receiving 5 times of chemotherapy, the condition was relieved, but the blood cells remained low, accompanied by complications such as susceptibility to infection and physical discomfort. After many medical consultations, he received CIK immune cell therapy in July 2018, but with little effect. In August 2018, he contacted the applicant and received the first course of treatment on September 8, 2018 (one course of treatment: one course of treatment) A bottle of PD-1-T cell preparation contains 2.5-3 billion PD-1-T cells, a bottle of PD-1-T cell preparation is injected every day, and 7 consecutive days is a course of treatment) PD-1-T cell therapy. The results before and after treatment are as follows.

[0099] On...

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Abstract

An embodiment of the invention relates to the technical field of immunotherapy and discloses a PD-1 antibody loaded T cell in-vitro culture method, a cell preparation and application. Specifically, the culture method includes steps: performing first-time culture by mixing peripheral blood mononuclear cells with an A-type culture medium; then, performing second-time culture by mixing a product obtained in first-time culture with a B-type culture medium; finally, mixing a product obtained after second-time culture with a K-type culture medium. By the culture method, lymphocytes separated from whole blood can be proliferated and differentiated into CD8+T cells in a culture process, the growing rate of separated T cells reaches 40 times or more within 18 days, and the CD8+T cell ratio exceeds80%.

Description

technical field [0001] The present invention relates to the technical field of immunotherapy, in particular, to an in vitro culture method of T cells loaded with PD-1 antibody, its cell preparation and its application. Background technique [0002] Today's cancer still fails to overcome the disease problem. In 2013, immunotherapy was hailed by the "Science" magazine as the most likely way to cure cancer. Immune cell therapy is the key part of immunotherapy. Immune cell therapy started in the 1970s, and the earliest was LAK cells (extracting peripheral blood from the human body, separating lymphocytes from peripheral blood in vitro, adding large doses of interleukin-2 Carry out in vitro culture and proliferation, and then obtain lymphocytes that proliferate several times and name them as LAK cells, and then make the cultured LAK cells into a cell suspension preparation and inject them into the human body intravenously). LAK cells are the earliest immune cells that are used in...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0783
Inventor 麦志国谢新波
Owner 广州筑康生物技术有限公司
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