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Application of HES6 serving as molecular target in treatment of chronic myeloid leukemia

A technology for chronic myeloid and leukemia, applied in the fields of biomedicine and clinical medicine

Active Publication Date: 2017-12-12
CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The function of HES6 in chronic myeloid leukemia (CML) and erythroid differentiation has not been reported at home and abroad

Method used

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  • Application of HES6 serving as molecular target in treatment of chronic myeloid leukemia
  • Application of HES6 serving as molecular target in treatment of chronic myeloid leukemia
  • Application of HES6 serving as molecular target in treatment of chronic myeloid leukemia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1: Packaged knockdown virus knocks down HES6 in hematopoietic stem cells

[0034] 1. To package HES6 overexpression / knockdown virus particles in 293T cells, first perform cell transfection according to the following system (Table 1):

[0035] Table 1. Plasmid transfection system reference table

[0036]

[0037] After 6 hours of transfection, change to fresh complete medium and time it; collect the virus stocks at 24 hours and 48 hours respectively, concentrate by ultracentrifugation, and store at -80°C after measuring the virus titer.

[0038] 2. Infect the packaged HES6 knockdown virus and control virus into K562 cells, and carry out the infection according to the conditions of MOI=50 and polybrene final concentration of 8ug / mL.

[0039] 3. Collect the cells to extract the total protein, and detect the expression of HES6 by Western blot. The specific method is: use RIPA protein lysate (P0013B, Biyuntian) to extract protein, and use BCA kit to measure prot...

Embodiment 2

[0042] Example 2: Expression of HES6 in newly diagnosed patients with CML

[0043] 1. Collect the mononuclear cells in the peripheral blood samples of normal people and newly diagnosed CML patients, use lymphocyte separation medium to separate the mononuclear cells in the peripheral blood samples of newly diagnosed CML patients and normal people as controls, and extract the total RNA of the cells for subsequent experiments .

[0044] 2. Use the real time PCR method to detect the expression of HES6, using GAPDH as the internal control: treat the mononuclear cells obtained in the sample in step 1 according to the TRIzol reagent provided by Invitrogen, extract the total RNA, and obtain cDNA by reverse transcription. HES6 and GAPDH primer sequences (see Table 2), and real time PCR was carried out according to the corresponding reaction system (see Table 3) and reaction conditions (see Table 4), to detect the expression of HES6 gene in newly diagnosed CML patients and normal contro...

Embodiment 3

[0052] Example 3: Packaged overexpression virus overexpresses HES6 in K562 cells

[0053] 1. To package HES6 overexpression / knockdown virus particles in 293T cells, first perform cell transfection according to the above-mentioned system in Table 1.

[0054] After 6 hours of transfection, change to fresh complete medium and time it; collect the virus stocks at 24 hours and 48 hours respectively, concentrate by ultracentrifugation, and store at -80°C after measuring the virus titer.

[0055] 2. Infect the packaged HES6 overexpression / knockdown virus and control virus into K562 cells, and infect according to the conditions of MOI=50 and final polybrene concentration of 8ug / mL.

[0056] 3. Collect the cells to extract the total protein, and detect the expression of HES6 by Western blot. The specific method is: use RIPA protein lysate (P0013B, Biyuntian) to extract protein, and use BCA kit to measure protein concentration. Immunoblotting was performed using the Westernblot method...

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PUM

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Abstract

The invention provides a novel molecular target HES6 for treating chronic myeloid leukemia (CML), correspondingly provides a HES6-based early diagnosis kit for treating the chronic myeloid leukemia, and further provides a method for promoting K562 cell differentiation through HES6 overexpression.

Description

technical field [0001] The invention belongs to the technical field of biomedicine and clinical medicine, and relates to a new therapeutic agent related to the treatment of chronic myelogenous leukemia and an application method thereof. Background technique [0002] HES6, located on human chromosome 2q37.3, is a member of the HES transcription family. As a transcriptional regulator, HES6 is expressed in many tissues, and it is involved in the regulation of neuron generation, myocyte generation, and intestinal tissue development. learning process. 成熟的HES6蛋白含224个氨基酸,其序列为:MAPPAAPGRDRVGREDEDGWETRGDRKARKPLVEKKRRARINESLQELRLLLAGAEVQAKLENAEVLELTVRRVQGVLRGRAREREQLQAEASERFAAGYIQCMHEVHTFVSTCQAIDATVAAELLNHLLESMPLREGSSFQDLLGDALAGPPRAPGRSGWPAGGAPGSPIPSPPGPGDDLCSDLEEAPEAELSQAPAEGPDLVRAALGA VTTAQIARSVWRPW。 [0003] HES6 is related to prostate cancer, breast cancer, metastatic colon cancer, malignant glioma and other cancers. Current research reports that HES6 can assist in the diagnosis of...

Claims

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Application Information

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IPC IPC(8): C12Q1/68A61K45/00A61P35/02
CPCA61K45/00C12Q1/6886C12Q2600/158C12Q2600/166
Inventor 刘静韩旭陈潇张洁莹张济张毅彬
Owner CENT SOUTH UNIV
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