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Double-gene modified stem cell and use thereof

A technology of mesenchymal stem cells and amino acids, which is applied in the preparation of drugs for the treatment of metabolic diseases in subjects, in the field of treatment of metabolic diseases, and can solve the problems of not being able to fundamentally improve the cause of the disease, long-term medication, etc.

Active Publication Date: 2019-06-25
BEIJING JIYUAN BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the clinical application of GLP1-Fc and FGF21, patients need to face long-term medication, life-long medication, and combined medication to lower blood sugar and lipids, and these drugs can only have the effect of treating the disease, and cannot fundamentally improve the cause of the disease

Method used

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  • Double-gene modified stem cell and use thereof
  • Double-gene modified stem cell and use thereof
  • Double-gene modified stem cell and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0158] Example 1. Preparation of FGF21 / GLP1-Fc Modified Mesenchymal Stem Cells

[0159] 1.1 Isolation and culture of autologous adipose-derived stem cells

[0160] Adipose derived mesenchymal stem cells (AD-MSCs) were isolated and cultured by mixed collagenase digestion method, the specific method is as follows:

[0161] Transfer the healthy adult adipose tissue extracted by liposuction to a 50mL centrifuge tube, add PBS for sufficient washing, and centrifuge at 1500rpm for 5 minutes to obtain the upper layer of adipose tissue. According to the ratio of 1:1:1, type I, II and IV collagenases were mixed to prepare 0.2% mixed collagenase, and the adipose tissue was added into the mixed collagenase digestion solution according to the ratio of adipose tissue: collagenase=1:1, Digest adipose tissue in a shaker at 37°C for 30 minutes. The digested adipose tissue was immediately added to 10% FBS α-MEM cell culture medium (purchased from Gibco), centrifuged at 1500 rpm for 10 minut...

Embodiment 2

[0177] Example 2. In vitro biological activity evaluation of FGF21 / GLP1-Fc modified mesenchymal stem cells

[0178] 2.1 Effect of MSC-FG on glucose-stimulated insulin secretion

[0179] Pre-preparation: Culture MSC, MSC-FG, MSC-FGF21 and MSC-GLP1-Fc cells obtained in Example 1 respectively in 100mm culture, when the confluence of the cells reaches 70%-80%, discard the original MSC serum-free medium, 10ml α-MEM medium, 37°C, 5% CO 2 Saturated humidity continued to cultivate for 48h. The culture supernatants of the four kinds of cells were collected, concentrated 10 times with an ultrafiltration column, and stored at 4°C for future use. Long-term storage needs to be placed in a -80°C refrigerator.

[0180] Take out a tube of frozen INS-1 cells (rat insulinoma cells, donated by the Academy of Military Medical Sciences) from liquid nitrogen and quickly put it in a 37°C water bath until the ice cubes disappear, and add 5 ml of preheated medium dropwise In a 15ml centrifuge tu...

Embodiment 3

[0189] Example 3. In vivo biological activity evaluation of FGF21 / GLP1-Fc modified mesenchymal stem cells

[0190] 3.1 Experimental grouping

[0191] Thirty 5-week-old male diabetic model mice BKS.Cg-Dock7m+ / +Leprdb / Nju (purchased from the Institute of Model Animals, Nanjing University) were selected. The experiment was divided into: control group (normal saline), liraglutide drug group (drug), MSC group (cell), MSC-FGF21 group (cell), MSC-FG group (cell) and MSC-GLP1-Fc group ( cells), a total of 6 groups, 5 mice in each group.

[0192] 3.2 Treatment plan

[0193] The administration method of liraglutide drug group (Liraglutide) was subcutaneous injection, and the administration method of the other groups was intraperitoneal injection, and all groups required administration in the morning.

[0194] Dosing time: The cell group requires that the cells be injected every 7 days, and the cells are given 3 times in total. The liraglutide drug group required the drug to be given t...

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Abstract

The invention relates to the field of cell therapy. In particular, the present invention relates to modified mesenchymal stem cells and a culture supernatant thereof, and a pharmaceutical compositioncomprising the cells or the culture supernatant thereof. The invention also relates to the use of the modified mesenchymal stem cells and the culture supernatant for treatment of metabolic disorder ina subject (eg, a human) and the use of the modified mesenchymal stem cells in preparation of a medicine for treating the metabolic disorder in the subject (eg, a human). The invention further relatesto a method of treating the metabolic disorder, which comprises the step of administering to the subject in need thereof the modified mesenchymal stem cell or the culture supernatant or the pharmaceutical composition thereof.

Description

technical field [0001] The present invention relates to the field of cell therapy. Specifically, the present invention relates to a modified mesenchymal stem cell and its culture supernatant, and a pharmaceutical composition comprising such cells or its culture supernatant. The present invention also relates to the use of the modified mesenchymal stem cells and their culture supernatant for treating metabolic disorders in a subject (for example, a human), and for preparing them for use in a subject (for example, a human) ) for use in a medicament for the treatment of metabolic disorders. The present invention also relates to a method for treating metabolic disorders, which includes the step of administering the modified mesenchymal stem cells of the present invention or their culture supernatant or pharmaceutical composition to a subject in need. Background technique [0002] Metabolic disorders such as diabetes are closely related to some endogenous molecules involved in ...

Claims

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Application Information

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IPC IPC(8): C12N5/10C07K14/50C07K14/605C07K19/00A61P3/04A61P3/10A61P3/06A61P1/16A61P3/00A61P9/10A61P9/12A61P27/02A61P25/00A61P13/12
CPCC12N5/0662C12N5/0667C07K14/50C07K14/605A61K35/28A61K38/1825A61K38/26A61K48/005A61K48/0008A61K45/06A61P3/10A61P3/06A61P1/16A61P5/50A61P3/00A61P9/10A61P9/12C12N2510/02C07K2319/30C07K2319/31A61K2300/00A61K48/00A61P3/04A61P13/12A61P25/00A61P27/02C07K19/00C12N2510/00C12N2501/119A61K38/00
Inventor 段海峰薛冰华谢晶张振利
Owner BEIJING JIYUAN BIOTECH
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