Application of GSK503 in preparing drug for treating RB (Retinoblastoma)

A technology of 1. GSK503 and retina, which is applied in the field of retinoblastoma treatment, can solve the problems of unclear curative effect and mechanism of action of retinoblastoma, and there is no GSK503 yet

Inactive Publication Date: 2017-05-24
SHANGHAI NINTH PEOPLES HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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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 the application of GSK503 in retinoblast...

Method used

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  • Application of GSK503 in preparing drug for treating RB (Retinoblastoma)
  • Application of GSK503 in preparing drug for treating RB (Retinoblastoma)
  • Application of GSK503 in preparing drug for treating RB (Retinoblastoma)

Examples

Experimental program
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Effect test

Embodiment 1

[0018] Immunofluorescence experiment

[0019] Experimental materials: retinoblastoma tissue chip (including 28 cases of retinoblastoma tissue and 12 cases of normal eye tissue), purchased from Xi'an Aili Biotechnology Co., Ltd.; H3K27me3 antibody and secondary antibody, purchased from Abcam (Abcam (Shanghai) Trading Co., Ltd.).

[0020] Place the tissue chip in a wet box, and spread PBS on the tissue chip. The diluted H3K27me3 antibody was centrifuged at 13500g for 2min at 4°C. Aspirate the PBS buffer on the glass slide at one end of the tissue chip, and add the antibody from the other end, cover with a humidification box, and incubate at room temperature for 1 h. Wash slides with PBS 3 times (5min / time), add new PBS buffer from one end of the slice, and absorb old buffer from the other end. The diluted secondary antibody was centrifuged at 13500 g for 2 min at 4°C. Add the secondary antibody on the tissue chip, incubate for 1 h at room temperature in a humidified box, and...

Embodiment 2

[0022] Chromatin immunoprecipitation (ChIP) experiments

[0023] Experimental materials: human retinoblastoma cell line RB44 cells, 37°C, 5% CO 2 Conventional culture, the medium was RPMI-1640 (Gibco) with 10% fetal bovine serum (FBS); GSK503 was purchased from MedChemExpress (MCE China); ChIP kit was purchased from millipore (Millipore, USA); PCR kit was purchased from Takara ( Treasure Bioengineering (Dalian) Co., Ltd.)

[0024] RB44 control cells and GS503-treated cells (10 μM GSK503, 37°C, 5% CO 2 Conventional culture for 7 days) centrifuge at 800g for 5min, discard the supernatant, resuspend in 10ml of 10% FBS 1640 culture medium, add 270μl of 37% formalin, incubate at room temperature for 10min, add 600μl of 2M glycine to stop the reaction, centrifuge at 4°C, 2500g for 15min , Discard the supernatant and lyse with cell lysate. Sonicate for 5 minutes to break up the DNA, centrifuge at 13500g for 15 minutes at 4°C, take the supernatant and perform ChIP experiments accor...

Embodiment 3

[0026] Cell Proliferation Assay (CCK8)

[0027] Experimental materials: CCK8 was purchased from Japan Tongren Chemical Co., Ltd.

[0028] RB44 cells were incubated with 10% FBS 1640 medium containing GSK503 at a final concentration of 10 μM at 37°C in 5% CO 2 Routine culture for 7 days, inoculated the treated cells and control RB4 cells in 96-well plate, 5000 cells per well, 100ul culture medium, 37°C, 5% CO 2 For culture, add 10ul CCK8 at 0h, 24h, 48h, and 72h respectively, continue to incubate at 37°C for 4h, and measure the absorbance value on the machine (see image 3 ). from image 3 It can be seen that GSK503 can significantly inhibit the proliferation of retinoblastoma, and the inhibitory effect is enhanced with the increase of concentration.

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Abstract

The invention belongs to the field of treating RB (Retinoblastoma), and particularly relates to application of GSK503 in preparing a drug for treating the RB. In the drug, the final concentration of the GSK503 is greater than or equal to 1 mu M. The GSK503 is a novel EZH2 (Enhancer of Zeste Homolog 2) small-molecule inhibitor, and an applicant experimentally finds that the GSK503 is capable of remarkably inhibiting the growth of the RB, and the methylation degree of H3K27me3 in cells is reduced through inhibiting the EZH2, so that cancer suppressor genes can be in re-expression, and the effect of inhibiting the tumor growth is reached. Effective treatment concentration can be reached when the final concentration of the GSK503 reaches 1 mu M, the use effect of the drug is increased while the concentration is increased, and the growth of the RB can be remarkably inhibited.

Description

technical field [0001] The invention belongs to the field of retinoblastoma treatment, and particularly relates to the application of GSK503 in the preparation of drugs for treating retinoblastoma. Background technique [0002] Retinoblastoma (RB) is the most common intraocular malignant tumor in children. It often develops during development. It is an embryonal malignant tumor originating from the nuclear layer of the retina. The tumor has a family genetic tendency, and is the most serious and harmful malignant tumor among infant eye diseases. Its visual prognosis is poor, and it is prone to intracranial and distant metastasis, which often endangers the lives of patients. [0003] At present, the treatment for this tumor is mainly non-etiological treatment such as surgery, radiotherapy and chemotherapy, and there is a lack of treatment methods for its etiology. Enhancer of Zeste gene homolog 2 (EZH2) is a member of the polycomb complex (PcG) family and is the catalytically...

Claims

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Application Information

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IPC IPC(8): A61K31/496A61P35/00
CPCA61K31/496
Inventor 范先群张赫文旭洋柴佩韦葛盛芳贾仁兵
Owner SHANGHAI NINTH PEOPLES HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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