Application of miR-31-5p in acute myelogenous leukemia

A pre-mir-31, acute myeloid technology, applied in the application field of miR-31-5p in the diagnosis and treatment of acute myeloid leukemia, can solve the problem of increased relapse rate of leukemia patients, and achieve the effect of prolonging life

Pending Publication Date: 2021-11-02
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Whether in long-term cell culture or in animal models, LSCs can cause and maintain leukemia; at the same time, unlike leukemia cells, more than 95% of LSCs are in the "dormant" state of G0 phase, and these cells are not effective for traditional cell cycle chemotherapy. Drug insensitivity, retained LSC increases the relapse rate of leukemia patients through self-replication (Renewal)

Method used

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  • Application of miR-31-5p in acute myelogenous leukemia
  • Application of miR-31-5p in acute myelogenous leukemia
  • Application of miR-31-5p in acute myelogenous leukemia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Example 1, a clinical sample collection and cell sorting embodiment

[0070] 1, collection of clinical samples AML

[0071] Hematology from acute myeloid leukemia random hospitalization (AML) patients with newly diagnosed 44 cases of bone marrow (n = 44), and bone marrow samples from patients were collected, and the collected sample number (AML-1, AML-2 , ......, AML-44), the patient did not receive any drug therapy; normal human bone marrow samples were collected 5 cases (n = 5). All the acquired samples were agreed by the ethics committee of the organization.

[0072] Collecting bone marrow sample processing method, and normal human bone marrow AML patient samples as follows:

[0073] 2mL marrow under sterile conditions, heparin was added. Bone marrow samples collection requirements: heparin, no blood clots, bone marrow greater than 2mL, samples were collected for cell sorting within 24 hours.

[0074] 2, isolated mononuclear cells

[0075] Preferred use of the company wh...

Embodiment 2

[0094] 2 detection, gene expression in miR-31-5p Example

[0095] 1, total RNA was extracted:

[0096] A, Example 1 points selected mononuclear cells or CD34 + CD38 - Stem cells were washed twice with PBS.

[0097] B, 1mL Trizol lysis buffer was added, mixed by pipetting several cleavage; 5min standing at room temperature so that adequate separation nucleoprotein.

[0098] C, and 200μL of chloroform, the tube cap tightly, shake vigorously 15s, room temperature for 5min.

[0099] D, 4 ℃, 12,000g, centrifuged 15min, learn about 400μL upper aqueous phase to another new 1.5mL EP tube. Be careful not to draw intermediate interface layer.

[0100] E, 500μL of isopropanol was added, thoroughly mixed, left at room temperature 10min.

[0101] F, 4 ℃, 12,000g, centrifuged 10min, the supernatant discarded, RNA precipitate at the bottom of the centrifuge tube.

[0102] G, was added 1mL 70% RNA was washed with ethanol precipitation.

[0103] H, 4 ℃, 7,500g, centrifuged 5min, discarded the supe...

Embodiment 3

[0130] Example 3 Analysis of miR-31-5p prognosis gene of AML patients from the database TCGA

[0131] (Https: / / portal.gdc.cancer.gov / ) TCGA-LAML the original miRNA expression by GDCRNATool (http: / / bioconductor.org / packages / devel / bioc / html / GDCRNATools.html) R download language pack GDC data and clinical information. Replicates miRNA raw data filtering the combined miRNA raw count data into a single matrix expression. Voom normalization and conversion for TMM, by running gdcVoomNormalization function, the raw data will be normalized by the counting method edgeR TMM implemented, and further converted by the methods limma voom provided. R using the language pack GDCRNATools gdcSurvivalAnalysis toolkit Kaplan Meier (KM) analysis, introduced miR-31-5p TCGA expression data and clinical information database, by grouping the expression of miR-31-5p, in order to maximize Analysis and found that sensitivity with a preset cut-off value (e.g. median) independent of any potential correlation va...

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Abstract

The invention relates to application of miR-31-5p in acute myelogenous leukemia, in particular to application of a product for detecting the miR-31-5p in preparation of a reagent, a chip or a kit for auxiliary diagnosis of subjects suffering from acute myelogenous leukemia (AML) and/or prognosis of the lifetime of the subjects. The invention also relates to an application of the miR-31 initial/precursor miRNA (prime/pre-miR-31) and/or the miR-31 mature miRNA (miR-31-5p) in preparation of a medicine for treating AML (acute myeloid leukemia). The miR-31-5p is introduced into cells through a gene transduction method, so bone marrow AML cells and leukemia stem cells (AML-LSC) can be induced to die, and the cytotoxicity of the chemotherapeutic drug cytarabine is enhanced; by expressing the miR-31-5p, the growth of AML cells and AML-LSC cells can be inhibited, and the life of animals can be prolonged. The disclosure provides a new method for diagnosis and prognosis evaluation of AML, and also provides a gene therapy drug for preparing AML.

Description

Technical field [0001] The present disclosure is in the field of biomedicine, it relates to the diagnosis and treatment of tumors, particularly to the use of miR-31-5p in acute myeloid leukemia (AML) in diagnosis and treatment. Background technique [0002] Leukemia is a type of clonal origin of hematopoietic stem cells of malignant disease, according to statistics, currently there are about 35 million people worldwide died of leukemia each year. Therefore, leukemia cancer has become the main current threat to human health. Leukemic stem cells (1eukemia stem cells, LSC) are present in a patient group of a trace amount of leukemic cell population, about 0.1% to 1% of all leukemic cells. LSC was first found in acute myeloid leukemia (AML) and has been widely recognized. Whether the long-term cell culture or in vivo animal models, and to maintain the LSC can cause leukemia; simultaneously, with different leukemia cells, more than 95% of the G0 state LSC "sleep", these cells periodic...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/113A61K31/7105A61P35/00
CPCC12Q1/6886A61K31/7105A61P35/00C12Q2600/178C12Q2600/158
Inventor 闫道广钟文彬
Owner JINAN UNIVERSITY
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