Use of TNFSF15 protein in preparation of melanoma treatment medicine

A melanoma and protein technology, applied in the field of medicine, can solve the problems of difficult treatment of metastatic melanoma, poor prognosis of patients, and low average survival rate, and achieve short production cycle, convenient scale-up production, and improved stability and activity Effect

Inactive Publication Date: 2018-01-05
NANKAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, for existing treatment methods, metastatic melanoma is very stubborn and difficult to treat, and the prognosis of patients will be very poor. The average six-month and five-year survival rate of these patients is less than 5%.

Method used

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  • Use of TNFSF15 protein in preparation of melanoma treatment medicine
  • Use of TNFSF15 protein in preparation of melanoma treatment medicine
  • Use of TNFSF15 protein in preparation of melanoma treatment medicine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1: TNFSF15 inhibits the migration ability of mouse melanoma cell B16

[0049] a) B16 cells at 37°C, 5% CO 2 The culture was grown to the logarithmic phase under certain conditions, digested, counted, spread into 6-well plates, 5X 10 per well 5 cells, adhere to the wall for 12 hours, so that the fusion degree of the cells is close to 100%;

[0050]b) Use a yellow pipette tip perpendicular to the well plate to draw a line along the ruler, one line every 0.5cm in each well, three lines in each well, wash twice with PBS, and wash away unattached cells;

[0051] c) Add 2 mL of serum-free medium to each well, take pictures with 10X objective lens and 10X eyepiece under white light, and record the scratch width at 0h;

[0052] d) Add TNFSF15 at final concentrations of 0, 10, 25, and 50 μg / mL, at 37°C, 5% CO 2 Cultivate for 24h under the condition of;

[0053] e) Take a picture with a 10X objective lens and 10X eyepiece under white light, and record the scratch widt...

Embodiment 2

[0055] Example 2 TNFSF15 induces apoptosis of mouse melanoma cell B16

[0056] a) B16 cells at 37°C, 5% CO 2 The culture was grown to the logarithmic phase under certain conditions, digested, counted, spread into 12-well plates, 2.5X 10 per well 4 cells, adhere to the wall for 12h.

[0057] b) Add TNFSF15 at final concentrations of 0, 1, 5, and 10 μg / mL, at 37°C, 5% CO 2 Cultivate for 48 hours under the same conditions.

[0058] c) Extraction of total cell protein: remove the cell culture medium, quickly wash the cells 3 times with pre-cooled PBS; add 100 μL of pre-cooled protein lysis buffer (50mM Tris-HCl. NaCl, 1mM EDTA, 1mMPMSF), use a cell scraper to lyse the cells; collect the liquid mixture in a centrifuge tube, incubate on ice for 30 minutes; centrifuge at 20,000g for 30 minutes at 4°C; take the supernatant, use the BCA method to determine the protein concentration, and use the Western Blotting method Detect the protein level of Cleaved-Caspase 3 in cells.

[0059...

Embodiment 3

[0061] Example 3 TNFSF15 can inhibit the growth of B16 cells, and improve the inhibitory ability of cisplatin to the growth of B16 cells

[0062] a) B16 cells at 37°C, 5% CO 2 The culture was grown to the logarithmic phase under certain conditions, digested, spread into 96-well plates, 10 per well 3 cells, adhere to the wall for 12h.

[0063] b) DDP group / DDP+TNFSF15 group were added with different concentrations of DDP and TNFSF15 protein with a final concentration of 5 μg / mL. The specific concentration of DDP is shown in the table below, with 4 replicate wells for each concentration gradient.

[0064]

[0065] 37°C, 5% CO 2 Stimulate 72h under the conditions.

[0066] c) The medium in the well plate was discarded, washed twice with PBS, and serum-free medium containing a final concentration of 0.5 μg / mL MTT was added to each well. 37°C 5% CO 2 Incubate for 6h.

[0067] d) The medium containing MTT was discarded, washed twice with PBS, and 150 μL of DMSO was added t...

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Abstract

The invention relates to a use of a TNFSF15 protein in the preparation of a melanoma treatment medicine. It is found that the TNFSF15 can inhibit the growth of endothelial cells and induce the apoptosis of the endothelial cells, and also can inhibit the growth and migration of mouse melanoma cells B16 and induce the apoptosis of the mouse melanoma cells B16. When the TNFSF15 and a chemotherapeuticmedicine are simultaneously used to treat melanoma, the TNFSF15 can inhibit the generation of tumors by inhibiting the generation of tumor blood vessels, and also can enhance the killing effect of the chemotherapeutic medicine on melanoma cells by inducing melanoma cell apoptosis. The TNFSF15 can enhance the melanoma treatment effect of cisplatin when used in combination with the chemotherapeuticmedicine cisplatin which is commonly used to treat melanoma.

Description

technical field [0001] The invention relates to the field of medicine, in particular to the use of TNFSF15 protein in preparing medicine for treating melanoma, and its use in combination with cisplatin in preparing medicine for treating melanoma. Background technique [0002] Malignant melanoma (MM) is a tumor that occurs in melanocytes of the skin or other organs due to the joint influence of genetic factors and environmental factors. The clinical manifestations of this disease are tenderness, itching, bleeding, ulcers, etc. The symptoms of the elderly are more serious than those of the young. This malignancy is growing much faster than other types, and the incidence of melanoma in the United States has doubled in the past 30 years. The World Health Organization reports that 132,000 such malignancies are diagnosed worldwide each year. If malignant melanoma is diagnosed early, the tumor can be surgically removed, which is used in 80% of cases. However, for existing treatm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/19A61P35/00C07K14/525C07K1/36C07K1/34C07K1/16A61K33/24
Inventor 李鲁远张智松高珊
Owner NANKAI UNIV
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