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Use of histone methyltransferase inhibitors in the preparation of products that promote megakaryocyte proliferation or platelet production

A technology of methyltransferase and megakaryocytes, which is applied in the field of histone methyltransferase inhibitors and the preparation of products that promote the proliferation of megakaryocytes or platelet production, which can solve the problems of low platelet levels and achieve low application cost and operability Strong and reproducible, high platelet production efficiency

Active Publication Date: 2021-02-19
血源生物科技(天津)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] One aspect of the present invention is aimed at the lack of compounds and methods that simultaneously promote cell proliferation, megakaryotic differentiation and platelet production in the prior art, and the low level of the compounds in the prior art to promote platelet production provides a histone methyl Use of transferase inhibitors in the preparation of products that promote megakaryocyte proliferation or platelet production

Method used

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  • Use of histone methyltransferase inhibitors in the preparation of products that promote megakaryocyte proliferation or platelet production
  • Use of histone methyltransferase inhibitors in the preparation of products that promote megakaryocyte proliferation or platelet production
  • Use of histone methyltransferase inhibitors in the preparation of products that promote megakaryocyte proliferation or platelet production

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] 1) cord blood CD34 + isolation of cells

[0062] a. Pour the umbilical cord blood into a plasma bottle or 50ml centrifuge tube, follow V 脐带血 :V 分选buffer =1:1, add sorting buffer (recipe as follows), add 1 / 4 hydroxyethyl starch of the above mixing volume, mix well, and let stand for 45min at room temperature;

[0063] Sorting buffer formula: phosphate buffered saline (PBS), 0.5% bovine serum albumin (BSA), 0.4% 500mM ethylenediaminetetraacetic acid (EDTA), 1% penicillin-streptomycin mixture (P / S)

[0064] b. Transfer the supernatant to a 50ml centrifuge tube, centrifuge at 1500rpm for 10min at room temperature; discard the supernatant, add sorting buffer to resuspend the cells; take a 15ml centrifuge tube, add 4.5ml of Ficoll separation solution (V Ficoll :V 分选buffer =1:1), slowly add the above-mentioned resuspension to the Ficoll separation solution to avoid destroying the liquid level stratification, 1800rpm, 15min (the centrifuge is set to 0 liters and 0 drops); ...

experiment example 1

[0075] Experimental Example 1: Cell Proliferation Detection

[0076] The cells at each stage in Example 1 were counted, and different control groups were set at the same time.

[0077] a. The control group (DMSO) and the experimental group (EHMTi) were cultured under the same conditions, and the cells were counted every three days to detect the effect of small molecule compounds on cell proliferation.

[0078] b. The result is as figure 1 As shown, compared with the DMSO control group, the addition of histone methyltransferase inhibitor (A366) treatment on the 0-3rd or 3-6th day of culture can significantly promote cell proliferation, and the cell expansion fold is increased by 15 times. above.

[0079] figure 1 On the 9th to 18th day of megakaryotic differentiation, the total number of cells in the control group (DMSO group) showed a downward trend; after adding histone methyltransferase inhibitor, the total number of cells still showed an upward trend on the 9th to 12th d...

experiment example 2

[0080] Experimental example 2: flow detection of megakaryocytes

[0081] The megakaryocytes in Example 1 were detected.

[0082] a. Collect about 1 × 10 on days 3, 6, 9, and 12 of megakaryocytic induction 5 The cell suspension was centrifuged at 300 g for 5 min, and the cell pellet was resuspended with 100 μL of 0.2% BSA.

[0083] b. Add 1 μL anti-CD41a-APC (BD) and 1 μL anti-CD42b-PE (BD) flow antibody to each group, incubate in the dark for 30 minutes, and then detect CD41a by flow cytometry (FACS Canto II; BD Biosciences). + CD42b + The proportion of megakaryocytes.

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Abstract

The invention discloses an application of histone methyltransferase inhibitor to preparation of a product for promoting megakaryocyte proliferation or blood platelet formation. The histone methyltransferase inhibitor is a histone methyltransferase G9a inhibitor. The histone methyltransferase G9a inhibitor stated in the invention can be used for promoting single CD34+hematopoietic stem and progenitor cells to differentiate and form 400 CD41a+CD42b+platelet granules, and the platelet granules are increased by 10 times than the platelet granules of a control group; the thrombopoiesis efficiency is further higher than the general range in literature report obviously, and a foundation is laid for mass production of functional platelets for clinic treatment in the future.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the use of a histone methyltransferase inhibitor in the preparation of a product for promoting megakaryocyte proliferation or platelet production. Background technique [0002] Platelets are one of the formed components of blood, and play an important role in the process of hemostasis and thrombosis in the body through functions such as adhesion, aggregation, and particle release. Clinically, various etiologies, such as hematopoietic malignancies, aplastic anemia, malignant tumors, hematopoietic stem cell transplantation, and radiotherapy and chemotherapy, may lead to thrombocytopenia or dysfunction, resulting in different degrees of bleeding in the body, and even endangerment in severe cases. life. Donor-derived platelet concentrate transfusion is currently the only clinical treatment. According to statistics, the number of patients who need platelet transfusions in China is nearl...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0786C12N5/078A61K35/19A61K35/28A61P7/06A61P7/04A61P1/16
CPCA61K35/19A61K35/28A61P1/16A61P7/04A61P7/06C12N5/0644C12N5/0645C12N2500/90C12N2501/125C12N2501/145C12N2501/2303C12N2501/72C12N2506/11
Inventor 周家喜刘懿莹刘翠翠苏培王洪涛
Owner 血源生物科技(天津)有限公司
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