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Diagnosis and treatment integrated apoptotic body and preparation method thereof

An apoptotic body and apoptotic cell technology, applied in the field of apoptotic bodies integrated with diagnosis and treatment and their preparation, can solve the problem of changing the conformation and biological activity of ligand molecules, reducing the recognition of tumor cells, and affecting the pharmacokinetics of probes. Learning and other problems to achieve the effect of prolonging the circulation time in the body

Active Publication Date: 2021-10-15
SHENZHEN INST OF ADVANCED TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, nano-drugs have been successfully verified in the precise diagnosis of tumors, and have shown good prospects for clinical application, but there are still some problems: chemical modification can easily change the conformation and biological activity of ligand molecules, and affect the drug resistance of probes. Kinetics, which reduces the ability of the probe to recognize tumor cells in a "contact" manner; therefore, it is urgent to develop new drugs that efficiently target and recognize tumors and use them for precise diagnosis and treatment of tumors

Method used

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  • Diagnosis and treatment integrated apoptotic body and preparation method thereof
  • Diagnosis and treatment integrated apoptotic body and preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0025] Example 1: Preparation of apoptotic bodies integrating diagnosis and treatment

[0026] (1) Macrophages were treated with antineoplastic drugs (doxorubicin) and nano-contrast agents (Fe 3 o 4 ) culture medium for 1-48h, the culture medium for apoptotic cells is obtained due to the toxicity of antitumor drugs inducing cell apoptosis;

[0027] (2) Collect the medium of apoptotic cells in a 50ml centrifuge tube and centrifuge at 900g for 10min to remove dead cells and cell debris. After centrifugation, in order to remove residual mixed cells in the supernatant, take the supernatant again and repeat Centrifuge once;

[0028] (3) Slowly move the supernatant to a 10ml sterile syringe, install a filter membrane filter device, apply slight pressure, and slowly pass the supernatant through a 5 μm filter membrane;

[0029] (4) Collect the filtrate in a centrifuge tube, put it into a high-speed centrifuge and centrifuge at 12000g-180000g for 18min-23min at 4°C, and the obtained...

Embodiment 2

[0031] Example 2: Testing the targeting effect of the apoptotic bodies prepared in Example 1 on tumors

[0032] Collect tumor cells in the logarithmic growth phase, use 10.0 mL of serum-free matrix, centrifuge at 1500 rpm for 3 min, and wash 1-2 times continuously; finally mix with serum-free matrix, and quantify the number of tumor cells with a hemocytometer ( Average cell count of 4 middle squares × 10 4 is the number of cells / ml). Tumor cell density was adjusted to 5 x 10 using serum-free stroma 6 pcs / mL (at least 2×10 6 per mL), each mouse was subcutaneously inoculated with 100 μL (ie 5×10 5 tumor cells), the tumor can be used in about 10 days.

[0033] A certain amount of apoptotic bodies were injected into the tail vein (the cell membrane of the apoptotic bodies was stained with the cell membrane dye DiD) and the distribution of the apoptotic bodies was observed at different time points. The experimental results were as follows: figure 2 shown, from figure 2 It c...

Embodiment 3

[0034] Example 3: Detecting the anti-tumor effect of the apoptotic body prepared in Example 1

[0035] Collect tumor cells in the logarithmic growth phase, use 10.0 mL of serum-free matrix, centrifuge at 1500 rpm for 3 min, and wash 1-2 times continuously; finally mix with serum-free matrix, and quantify the number of tumor cells with a hemocytometer ( Average cell count of 4 middle squares × 10 4 is the number of cells / ml). Tumor cell density was adjusted to 5 x 10 using serum-free stroma 6 pcs / mL (at least 2×10 6 per mL), each mouse was subcutaneously inoculated with 100 μL (ie 5×10 5 tumor cells), the tumor can be used in about 10 days.

[0036] 24 C6 glioma subcutaneous tumor nude mice were randomly divided into 4 groups: (1) apoptotic body, (2) control, (3) blank apoptotic body (apoptotic body induced by ultraviolet light), (4) ) Adriamycin group. 6 in each group. Wait until the volume of the subcutaneous tumor reaches 50mm 2 At the same time, a certain amount of ...

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Abstract

The invention discloses a diagnosis and treatment integrated apoptotic body and a preparation method thereof. The preparation method comprises the following steps: culturing cells in a culture medium containing an antitumor drug and a nano contrast agent to obtain a culture medium of apoptotic cells; centrifuging the culture medium of apoptotic cells to obtain supernate; filtering the supernate to obtain filtrate; and centrifuging the filtrate to obtain a precipitate which is the diagnosis and treatment integrated apoptotic body. According to the invention, a nano-probe is carried on the mononuclear cells by utilizing the characteristic that the mononuclear cells in blood actively swallow apoptotic bodies; the in-vivo circulation time of the nano-probe is prolonged by actively avoiding interception and removal of a reticuloendothelial system through the mononuclear cells; and under recruitment of chemotactic factors released by tumor stem cells, the mononuclear cells carry anti-tumor drugs and contrast agents to actively permeate into a tumor infiltration area. The novel probe design thought gives full play to the unique advantages of autoimmune cells, and breaks through the traditional mode of 'contact type' recognition of targeted ligands and receptors.

Description

technical field [0001] The invention belongs to the technical field of materials, and in particular relates to an apoptotic body integrating diagnosis and treatment and a preparation method thereof. Background technique [0002] Cancer is also called a malignant tumor. It is very difficult to cure, and it is a time bomb growing in the body. It grows fast. Once it breaks out, it will inevitably endanger life and health. Malignant tumors have become one of the major public health problems that seriously threaten the health of the Chinese population, and the prevention and control situation is grim. According to the latest global cancer burden data in 2020 released by the International Cancer Research Center of the World Health Organization, there will be 19.29 million new cancer cases in the world in 2020, of which 4.57 million are new cancer cases in China, accounting for 23.7% of the world, ranking first in the world. In 2020, there will be 9.96 million cancer deaths worldw...

Claims

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Application Information

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IPC IPC(8): A61K45/00A61K31/704A61K47/46A61P35/00A61K49/18C12N5/078C12N5/0786C12N5/09
CPCA61K45/00A61K31/704A61K47/46A61P35/00A61K49/1896C12N5/0645C12N5/0634C12N5/0693C12N2501/06
Inventor 郑海荣盛宗海胡德红刘新
Owner SHENZHEN INST OF ADVANCED TECH
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