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Preparation and application of a car-nk cell sustained-release agent for ovarian cancer

A slow-release agent and ovarian cancer technology, applied in the field of biomedicine, can solve problems such as lack of in-depth exploration, poor effect of cell preparations, and limitations in the application of CAR-NK cells

Active Publication Date: 2020-11-13
广东美赛尔细胞生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method is simple to operate, but the cell preparation it prepares cannot make the cells survive well, resulting in poor cell preparation effect
[0007] At present, most of the preparation methods for CAR-NK cells are obtained through virus transfection, but there is still a lack of in-depth exploration on how to effectively apply the obtained cells, which limits the application of CAR-NK cells

Method used

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  • Preparation and application of a car-nk cell sustained-release agent for ovarian cancer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 A preparation method of CAR-NK cell sustained release agent for ovarian cancer

[0031] S1 Collect the peripheral blood of the patient, add 1.5 times the volume of phosphate buffered saline to dilute, put the diluted peripheral blood into a centrifuge tube containing Ficoll lymphocyte separation medium, and centrifuge at 700g for 25min, absorb the buffy coat at the interface between the upper plasma layer and Ficoll Layer, get buffy coat cells;

[0032] S2 Wash the buffy coat cells obtained in step S1 twice with phosphate buffered saline, centrifuge at 800 g for 10 min after the first wash, and centrifuge at 400 g for 10 min after the second wash to obtain peripheral blood mononuclear cells;

[0033] S3 resuspend the peripheral blood mononuclear cells obtained in step S2 with serum-free RPMI1640 culture medium, and adjust the cell concentration to 2×10 6 / mL to obtain a cell suspension;

[0034] S4 Mix the cell suspension obtained in step S3 with the inactivate...

Embodiment 2

[0038] Example 2 A preparation method of CAR-NK cell sustained release agent for ovarian cancer

[0039] S1 Collect the peripheral blood of the patient, add 1.5 times the volume of phosphate buffered saline to dilute, put the diluted peripheral blood into a centrifuge tube containing Ficoll lymphocyte separation medium, centrifuge at 600g for 30min, and absorb the buffy coat at the interface between the upper plasma layer and Ficoll Layer, get buffy coat cells;

[0040] S2 Wash the buffy coat cells obtained in step S1 twice with phosphate buffered saline, centrifuge at 800 g for 10 min after the first wash, and centrifuge at 400 g for 10 min after the second wash to obtain peripheral blood mononuclear cells;

[0041] S3 resuspend the peripheral blood mononuclear cells obtained in step S2 with serum-free RPMI1640 culture medium, and adjust the cell concentration to 1×10 6 / mL to obtain a cell suspension;

[0042] S4 Mix the cell suspension obtained in step S3 with the inactiva...

Embodiment 3

[0046] Example 3 A CAR-NK cell sustained-release agent for ovarian cancer

[0047] S1 Collect peripheral blood from the patient, add 1.5 times the volume of phosphate buffered saline to dilute, add the diluted peripheral blood to a centrifuge tube containing Ficoll lymphocyte separation medium, and centrifuge at 800g for 20min, absorb the buffy coat at the interface between the upper plasma layer and Ficoll Layer, get buffy coat cells;

[0048] S2 Wash the buffy coat cells obtained in step S1 twice with phosphate buffered saline, centrifuge at 800 g for 10 min after the first wash, and centrifuge at 400 g for 10 min after the second wash to obtain peripheral blood mononuclear cells;

[0049] S3 resuspend the peripheral blood mononuclear cells obtained in step S2 with serum-free RPMI1640 culture medium, and adjust the cell concentration to 3×10 6 / mL to obtain a cell suspension;

[0050] S4 Mix the cell suspension obtained in step S3 with the inactivated autologous plasma, the...

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Abstract

Belonging to the biomedical field, the invention in particular relates to preparation and application of a CAR-NK cell sustained-release agent for ovarian cancer. The CAR-NK cell sustained-release agent is prepared by the method of: collecting the peripheral blood of patients, conducting washing and centrifugation to obtain a peripheral blood mononuclear cell, culturing the peripheral blood mononuclear cell for 7d to obtain NK cells, and conducting transfection amplification with BUHZOU lentivirus to obtain CAR-NK cells; at the same time preparing interleukin-2 nanoparticles; and finally, mixing a CAR-NK cell suspension, the interleukin-2 nanoparticles, a glucose solution, a sodium danshensu aqueous solution, a glutamine aqueous solution and a Pluronic F127 solution to obtain the CAR-NK cell sustained-release agent. The preparation method of the CAR-NK cell sustained-release agent provided by the invention is simple, the prepared sustained-release agent is more pure, immunological rejection does not occur easily, at the same time, cells in the sustained-release agent have good dispersibility, thus avoiding cell reunion, and significantly improving the killing activity of cancer cells.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to the preparation and application of a CAR-NK cell slow-release agent for ovarian cancer. Background technique [0002] Ovarian cancer (Ovarian cancer, OC) is one of the three major malignant tumors of the female reproductive system, and its mortality rate accounts for the second among gynecological malignancies. Ovarian cancer has become a malignant disease that seriously threatens the lives of patients due to its insidious onset, multiple histological types, lack of early diagnosis methods, and poor efficacy of surgery and radiotherapy. The five-year survival rate of ovarian cancer patients is low. [0003] Although traditional surgery, chemotherapy, and radiotherapy for OC can alleviate the condition, more than 60% of patients still experience tumor recurrence and metastasis after initial treatment. As a kind of tumor biotherapy, immunotherapy has shown great potential no...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K35/17A61K38/20A61K47/10A61P35/00C12N5/10
CPCA61K35/17A61K38/2013A61K47/10A61P35/00C07K14/7051C12N5/0646C12N2501/11C12N2501/2302C12N2501/2315C12N2501/2321C12N2501/599C12N2510/00A61K2300/00
Inventor 陈素红梅建勋徐单单
Owner 广东美赛尔细胞生物科技有限公司
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