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Application of Optineurin (OPTN) in diagnosis and treatment of ocular melanoma

A technology for melanoma and melanoma cells, applied in the application field of Optineurin in the diagnosis and treatment of ocular melanoma, can solve the problems of lack of in-depth research, and achieve the effect of inhibiting malignant progression and good clinical application value

Active Publication Date: 2021-08-31
SHANGHAI NINTH PEOPLES HOSPITAL SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no research on the relationship between OPTN and ocular melanoma in this field, and there is a lack of in-depth research on the relationship between OPTN and the occurrence and development of ocular melanoma, as well as the potential mechanism of its function in ocular melanoma

Method used

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  • Application of Optineurin (OPTN) in diagnosis and treatment of ocular melanoma
  • Application of Optineurin (OPTN) in diagnosis and treatment of ocular melanoma
  • Application of Optineurin (OPTN) in diagnosis and treatment of ocular melanoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Example 1 Immunofluorescent staining of tissue sections

[0023] Experimental materials: The tissue samples in this example come from ocular melanoma samples and ocular melanoma samples from the Department of Ophthalmology, Ninth People's Hospital Affiliated to Shanghai Jiaotong University School of Medicine. The study protocol was approved by the Hospital Ethics Committee, and the patients obtained written informed consent.

[0024] Experimental steps: 4% paraformaldehyde was used to fix human ocular melanoma tissue or melanoma tissue samples. Paraffin-embedded tissue sections were 5 μm thick and mounted on glass slides, and 1 mM EDTA was used for antigen retrieval. The sections were incubated in methanol containing 3% hydrogen peroxide to inactivate endogenous peroxidase activity, and then the sections were rinsed in PBS for 6 min. Sections were blocked in goat serum for 2 hours at room temperature, and then incubated overnight at 4°C with primary antibodies. Rinse...

Embodiment 2

[0025] Knockout of embodiment 2OPTN and plate colony formation experiment

[0026] Experimental materials: human uveal melanoma cell line MUM2B, human conjunctival melanoma cell line CRMM1. Lipofectamine 2000 was purchased from Thermo Fisher (USA), and 4% paraformaldehyde and crystal violet dye were purchased from Shenggong Company (China).

[0027] Experimental steps: (1) MUM2B, MEL290, CRMM1, CM2005.1, and PIG1 cells were conventionally cultured in a 5% CO2 incubator at 37°C. MUM2B and MEL290 were cultured in DMEM medium containing 10% FBS, and CRMM1, CM2005.1 and PIG1 were cultured in F12-K containing 10% FBS.

[0028] (2) Digest and centrifuge MUM2B and CRMM1 cells, resuspend them with complete medium, count 300,000 cells / well and spread them on a six-well plate. When the cells adhere to the wall and the growth density is 50%-60%, replace it with serum-free culture base.

[0029] (3) Set knockout group (siOPTN) and control group (siNC).

[0030] siOPTN-1:

[0031] sen...

Embodiment 3

[0038]Example 3 Knockout of OPTN and Transwell Cell Migration Experiment

[0039] Experimental materials: human uveal melanoma cell line MUM2B, human conjunctival melanoma cell line CRMM1. Lipofectamine 2000 was purchased from Thermo Fisher (USA), 8 μm 24-well plate Transwell chamber was purchased from Millipore (USA), 4% paraformaldehyde and crystal violet dye were purchased from Sangon (China).

[0040] Experimental steps: (1) (2) (3) The steps are the same as before.

[0041] (4) After identifying the knockout effect, the remaining cells were counted. 900 μl of 10% FBS medium was added to each well of the 24-well plate, 8 μm Transwell chambers were suspended in the wells, and 20,000 cells were inoculated in 250 μl of 2% FBS medium in each chamber. 37°C, 5% CO2 incubator after routine culture for 48h-72h, aspirate the culture medium in the well and chamber, carefully wash twice with PBS, fix with 4% paraformaldehyde for 30min, stain with crystal violet for 30min, carefully...

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Abstract

The invention provides application of OPTN in diagnosis and treatment of ocular melanoma. It is found that the expression level of the OPTN in the ocular melanoma is relatively high and is related to poor prognosis. In addition, the malignant progression of the ocular melanoma can be effectively inhibited by specifically knocking out the OPTN. Therefore, the OPTN can be used as a molecular biomarker for diagnosing and treating the ocular melanoma, and can be used as a prognosis index and a potential treatment target of the ocular melanoma.

Description

technical field [0001] The invention relates to the technical fields of molecular biology and medicine, in particular to the application of Optineurin in the diagnosis and treatment of ocular melanoma. Background technique [0002] Ocular melanoma includes uveal melanoma (Uveal melanoma, UM) and conjunctival melanoma (Conjunctival melanoma, CM). Among them, UM is the most common intraocular malignancy in adults, with a high degree of malignancy. About 50% of UM patients develop liver metastases, and the median survival time of patients with liver metastases is only 10.2 months. The onset of CM is relatively insidious, and often does not affect vision in the early stage. When it is found, it often spreads and metastasizes, and the prognosis is poor, which seriously affects the life and health of patients. At present, the main treatment methods for ocular melanoma are surgery and radiotherapy and chemotherapy, but it is not sensitive to conventional radiotherapy and chemother...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K45/00A61P35/00C12Q1/6886
CPCA61K45/00A61P35/00C12Q1/6886C12Q2600/118C12Q2600/158
Inventor 庄艾范先群柴佩韦王少云林惠民
Owner SHANGHAI NINTH PEOPLES HOSPITAL SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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