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Application of VCP (valosin containing protein) inhibitor and oncolytic virus in preparation of antitumor drugs

An oncolytic virus and inhibitor technology, applied in the field of biomedicine, can solve problems such as unclear mechanism of action

Active Publication Date: 2016-12-07
GUANGZHOU VIROTECH PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the patent 201510990705.7 previously applied by the inventor, chrysophanol and its derivatives are used as anti-tumor synergists of oncolytic viruses. The combination of the two can reduce the survival rate of tumor cells to 39.6%, but there is a big difference in its anti-cancer strength. room for improvement, in addition, the mechanism of action of this combined application is not yet clear

Method used

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  • Application of VCP (valosin containing protein) inhibitor and oncolytic virus in preparation of antitumor drugs
  • Application of VCP (valosin containing protein) inhibitor and oncolytic virus in preparation of antitumor drugs
  • Application of VCP (valosin containing protein) inhibitor and oncolytic virus in preparation of antitumor drugs

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Example 1 Eeyarestatin I and M1 virus significantly increase the morphological lesions of human liver cancer cell lines

[0072] Material:

[0073] Human hepatocellular carcinoma Hep3B, M1 virus, high glucose DMEM medium, inverted phase contrast microscope.

[0074] method:

[0075] a) Cell culture: Human hepatocellular carcinoma Hep3B was grown in DMEM complete medium containing 10% FBS, 100 U / ml penicillin and 0.1 mg / ml streptomycin; all cell lines were placed in 5% CO 2 , cultured and subcultured in a closed incubator with a constant temperature of 37°C (95% relative humidity), and observed the growth with an inverted microscope. The cells were subcultured every 2-3 days, and the cells in the logarithmic growth phase were taken for formal experiments.

[0076] b) Cell treatment and morphological observation: cells in the logarithmic growth phase were selected, and DMEM complete culture medium (containing 10% fetal bovine serum, 1% double antibody) was used to make...

Embodiment 2

[0079] Example 2 Combined treatment of Eeyarestatin I or CB-5083 with M1 virus significantly reduces the survival rate of human liver cancer cell lines

[0080] Material:

[0081] Human hepatocellular carcinoma Hep3B, M1 virus, high glucose DMEM medium, automatic enzyme-linked detection microplate reader.

[0082] method:

[0083] a) Cell inoculation, administration treatment: select logarithmic growth phase cells, DMEM complete culture medium (containing 10% fetal calf serum, 1% double antibody) to make cell suspension, with 4 × 10 3 The density per well was seeded in a 96-well culture plate. After 12 hours, the cells were completely attached to the wall. The experiment was divided into the control group without drug and virus treatment, the Eeyarestatin I or CB-5083 group alone, the M1 alone infection group and the Eeyarestatin I / M1 combination group or the CB-5083 / M1 combination group . The doses used were: the doses used were: M1 virus (MOI=0.001) infected cells; Eeyar...

Embodiment 3

[0090] Example 3 Knocking down the expression level of VCP can enhance the oncolytic effect of oncolytic virus M1 on liver cancer cells

[0091] Material:

[0092] M1 virus, human hepatocellular carcinoma Hep3B, human hepatocellular carcinoma Huh 7, VCP interference fragment (Si RNA), MTT (methyl azolazolium blue), Lipofectamine TM RNAiMAX (invertrogen, USA) Western bolt: cell total protein extract ( Mammalian Protein Extraction Reagent, Thermo), VCP antibody (CST, USA), GAPDH antibody (CST, USA);

[0093] method:

[0094] Select logarithmic growth phase cells, DMEM complete culture medium (10% fetal bovine serum, 1% double antibody) to make cell suspension, and the cells are 1 × 10 5 The density per well was seeded in a 6-well plate. After 24 hours, RNAiMAX-coated Si RNA fragments were added. After 48 hours, the M1 virus was infected. 48 hours after infection, samples were processed.

[0095] a) Add 20 μl (5 mg / ml) of MTT to each well, measure the absorbance value af...

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Abstract

The invention belongs to the field of biological medicine and relates to an application of a VCP (valosin containing protein) inhibitor and an oncolytic virus in preparation of antitumor drugs. The invention discovers that the VCP inhibitor can be used for preparing an oncolytic virus antitumor synergist. Meanwhile, the invention relates to pharmaceutical composition containing the VCP inhibitor and the oncolytic virus, a drug suit containing the VCP inhibitor and the oncolytic virus as well as an application of the VCP inhibitor and the oncolytic virus in treatment of tumors, especially tumors insensitive to the oncolytic virus. Besides, the invention relates to an antitumor medication system, and the system is characterized by comprising a VCP expression level detection reagent and the oncolytic virus.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to the application of the combination of a VCP protein inhibitor and an oncolytic virus in the preparation of antitumor drugs. Background technique [0002] Oncolytic virus is a kind of replication-competent virus that selectively infects and kills tumor cells without damaging normal cells. Oncolytic virus therapy (oncolytic virotherapy) is an innovative tumor-targeted treatment strategy, which uses natural or genetically engineered viruses to selectively infect tumor cells and replicate in tumor cells to achieve targeted dissolution, The effect of killing tumor cells, but no damage to normal cells. [0003] M1 virus (Alphavirus M1) belongs to the genus Alphavirus (Alphavirus), and it has a good application effect in the preparation of antitumor drugs. For example, the Chinese invention patent application 201410425510.3 discloses that the M1 virus can selectively cause tumor cell death wit...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K45/06A61K48/00A61K35/768A61P35/00A61K31/519
CPCA61K31/519A61K31/7088A61K35/768A61K45/06A61P35/00A61K31/4178A61K31/7105A61K2300/00A61K48/00A61K31/00C12N7/00C12N2770/36132A61K31/4439A61K31/713A61K38/1774A61K9/0019A61K9/19A61K9/48A61K9/7023
Inventor 张海鹏邓静林园W.K.凯夫尼程世源高光坪林穗珍吕凌蔡静龚守芳胡骏白雪涛肖晓李凯梁剑开谭亚倩朱文博银巍颜光美
Owner GUANGZHOU VIROTECH PHARMA
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