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SENP1 protein inhibitor and use thereof

An inhibitor and protein technology, applied in the field of SENP1 protein inhibitor and its application, can solve the problem that there is no target for the treatment of malignant hematological tumors

Inactive Publication Date: 2015-03-25
INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But so far, there is no report on using SENP1 as a therapeutic target for hematological malignancies.

Method used

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  • SENP1 protein inhibitor and use thereof
  • SENP1 protein inhibitor and use thereof
  • SENP1 protein inhibitor and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Example 1: Detection of SENP1 expression in CML.

[0084] The inventors performed magnetic bead sorting on the bone marrow cells of 8 CML patients and donors, and obtained CD34 + Cells, the expression of SENP1 mRNA was detected by real-time quantitative PCR, and it was found that the expression of SENP1 in CML patients was higher than that in donors ( figure 1 -A). Take two cases of CML CD34 + The cells were sorted for stem and progenitor cells to obtain myeloid progenitor cells (CMP), granulomonocytic progenitor cells (GMP), erythroid megakaryocytic progenitor cells (MEP) and hematopoietic stem cells (HSC), followed by PCR detected, and found that the SENP1 expression level of the HSC fraction was the highest ( figure 1 -B). The proteins of two samples were taken for western blot analysis, and it was found that the protein of SENP1 was in CML CD34 + cells than donor CD34 + The expression of cells should be high ( figure 1 -C).

Embodiment 2

[0085] Example 2: Construction and identification of shRNA (short hairpin RNA, small hairpin RNA) interference vector that reduces SENP1 gene expression.

[0086] Purchase 1 set (including 5 pieces) of SENP1 interference vector set (SHCLNG, NM-014554, including TRCN0000004395, TRCN0000004396, TRCN0000004397, TRCN0000004398, TRCN0000004399) from sigma company, and use lipofectin2000 transfection reagent to transfer engineering cell line to extract total HEK293 cells, RNA, and then use RT-qPCR detection to obtain the carrier with the highest interference efficiency. The specific process is: using the reverse-transcribed cDNA as a template, qPCR (quantitative PCR) to detect the expression of the SENP1 gene, using β-actin as an internal reference; The empty vector of the vector is used as a control, and 2-ΔΔCT is the expression of SENP1 after interference. The small hairpin sequence in the vector with the highest interference efficiency is: 5'CCGGGCGCCAGAUUGAAGAACAGAACUCGAGUUCUGUU...

Embodiment 3

[0088] Example 3: Preparation of lentivirus containing human SENP1-shRNA and the inhibitory effect of SENP1-GFP-shRNA on K562 cells.

[0089] After the successful construction of the shRNA interference vector that reduces the expression of the SENP1 gene, the lentivirus was prepared and purified. First inoculate 293T cells 4-4.5×10 6in a 10cm petri dish. The next day, the cells were confluent to 80-85%, and the lentiviral plasmid SENP1-GFP-shRNA was transfected. The lentiviral plasmid was transfected into 293T cells by the calcium phosphate transfection method, and the specific steps were: pMD2G: 3 μg, pSAX2: 9 μg (the vector was purchased from Addgene), and the target plasmid (interference vector): 12 μg. The three plasmids were mixed well. Add 63 μl of 2.0M CaCl 2 , fill up to 500μl with 0.1×TE, and mix well. Slowly drop the above liquid into 500μl 2×HBS, and mix well while adding. The liquid was in a turbid state after mixing, and stood at room temperature for 25-30mi...

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Abstract

The present invention relates to an SENP1 protein inhibitor and a use thereof, in particular, the invention relates to an SENP1 protein inhibitor shRNA, a use of the SENP1 protein inhibitor in preparation of drugs for preventing or treating malignant blood tumor, and a use in preparation of drugs for inhibiting the proliferation of malignant blood tumor cells and / or promoting the apoptosis of the malignant blood tumor cells. The invention further relates to a method for screening the drugs for preventing or treating malignant blood tumor.

Description

technical field [0001] The present invention relates to an inhibitor of SENP1 protein and its application, specifically, the present invention relates to an inhibitor shRNA of SENP1 protein, and the use of an inhibitor of SENP1 protein in the preparation of a drug for preventing or treating malignant hematological tumors, and for the preparation of Use in drugs for inhibiting the proliferation of malignant hematological tumor cells and / or promoting the apoptosis of malignant hematological tumor cells. The present invention also relates to methods of screening drugs for preventing or treating hematological malignancies. Background technique [0002] Hematological malignancies include leukemia, myeloma and other diseases. Chronic myeloid leukemia (CML) is a kind of malignant clonal blood disease mainly caused by malignant transformation of hematopoietic stem cells under the action of proto-oncoprotein BCR-ABL, also known as chronic myeloid leukemia. The pathogenesis of CML i...

Claims

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Application Information

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IPC IPC(8): A61K45/06A61K48/00C12N15/113C12N15/867C12N5/10C12Q1/68C12Q1/37G01N33/573A61P35/00A61P35/02
Inventor 王立生孙慧燕吴祖泽肖凤君王华杨月峰
Owner INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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