Autophagy blocking system based on nano materials, preparation method thereof and application in arsenical-drug solid-tumor treatment with same

A nanomaterial and autophagy technology, which is applied in the application field of arsenic drugs in the treatment of solid tumors, can solve the problems of unsatisfactory treatment effects of arsenic drugs, and achieve good medical application prospects, good therapeutic effects, and improved therapeutic effects. Effect

Active Publication Date: 2017-08-18
SHANGHAI INST OF APPLIED PHYSICS - CHINESE ACAD OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a nanomaterial-based autophagy blocking system and its preparation method, as well as its application in the treatment of solid tumors with arsenic drugs, so as to solve the unsatisfactory therapeutic effect of arsenic drugs on solid tumors in the prior art The problem

Method used

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  • Autophagy blocking system based on nano materials, preparation method thereof and application in arsenical-drug solid-tumor treatment with same
  • Autophagy blocking system based on nano materials, preparation method thereof and application in arsenical-drug solid-tumor treatment with same
  • Autophagy blocking system based on nano materials, preparation method thereof and application in arsenical-drug solid-tumor treatment with same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1: Comparison of the efficacy of ATO on acute promyelocytic leukemia cells NB4 and human liver cancer cells HepG2

[0047] ATO was purchased from Sigma (Cat. No.: 202673), dissolved in 0.1M NaOH solution, adjusted to pH 8 with 1M HCL, filtered and sterilized.

[0048] Human liver cancer cells HepG2 were purchased from Shanghai Cell Bank of Chinese Academy of Sciences and cultured in 1640 medium (Gibco) containing 10% fetal bovine serum. 37°C, 5% CO 2 , Saturated humidity cultivation. 7×10 4 / Well density was seeded in a 24-well plate and adhered overnight. Add ATO at concentrations of 2, 4, 8, 16, 32μM respectively, with cells without any treatment as a control, three replicate wells in each group, after 48h incubation, staining with MTT (Sigma) for 4h, adding 10% acidic SDS to dissolve it Formazan crystals, the ultraviolet absorption value of each hole was measured at OD 570nm, and the cell survival rate was expressed as the percentage of OD (treatment group) / OD (...

Embodiment 2

[0051] Example 2: Evaluation of the efficacy of ATO combined with autophagy blocking drug CQ on tumor cells.

[0052] CQ was purchased from Sigma (Cat. No.: C6628) and dissolved in sterile water.

[0053] In western blotting experiments, HepG2 cells were 7×10 4 The density of / well was inoculated in a 24-well plate and adhered overnight. Set up the following experimental groups, each with three replicates: CQ (12.5μg / mL), ATO (2μM and 4μM each), CQ+ATO mixture (12.5μg / mL CQ+2μM ATO and 12.5μg / mL CQ+ 4μM ATO each group), with the cells without any treatment as a control, incubate for 48h. Wash the cells twice with PBS, lyse the cells with 1×SDS loading buffer, denature at 95°C, transfer to PVDF membrane after polyacrylamide gel electrophoresis. The PVDF membrane was blocked with 6% skimmed milk powder dissolved in PBST (0.1% Tween 20) for 1 hour, washed with PBST 3 times, and a 1:1000 primary antibody was added. The primary antibodies involved are as follows: anti-LC3 (Novus), P6...

Embodiment 3

[0056] Example 3: Nano-diamond's autophagy regulation effect on cells.

[0057] Nanodiamonds (NDs, Gansu Jinshi Nanomaterials Co., Ltd.), the purity of the nanodiamonds> 99%, the single particle diameter is about 2-10nm, forming a cluster structure of about 250nm in the solution. Irradiate with ultraviolet for 30 minutes and disperse in sterile water.

[0058] In western blotting experiments, HepG2 cells were 7×10 4 The density per well was seeded in a 24-well plate and adhered overnight. 50μg / mL NDs and 12.5μg / mL CQ were added respectively, and the cells without any treatment were used as controls. Three replicate wells in each group were incubated for 48h. The methods of cell lysis, protein electrophoresis, and membrane transfer are the same as in the second embodiment. The primary antibodies involved in this example are as follows: anti-LC3 (Novus), P62 (Abcam) and GAPDH (Abcam), mTOR (cell signaling technology), P-mTOR (cell signaling technology), p70S6K (cell signaling techn...

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Abstract

The invention provides an autophapy blocking system based on nano materials, a preparation method thereof and application in arsenical-drug solid-tumor treatment with same. The preparation method comprises the following steps of dissolving arsenic trioxide in a NaOH solution, adjusting the pH value to 8-8.5, and obtaining an arsenic trioxide solution; dispersing nano materials in water or saline or a buffer system, and obtaining a nano material suspension; mixing the arsenic trioxide solution with the nano material suspension evenly, and obtaining the autophagy blocking system. The autophapy blocking system based on nano materials can effectively block a solid tumor autophagy induced by ATO, thereby significantly improving the killing effect of ATO on solid tumor cells so as to enable a low concentration ATO to have a good treating effect on the solid tumor and reduce the toxic and side effects on normal tissue.

Description

Technical field [0001] The invention relates to the field of biotechnology, and more specifically to an autophagy blocking system based on nanomaterials, a preparation method thereof, and application in the treatment of solid tumors with arsenic agents. Background technique [0002] Arsenic trioxide (ATO) is a traditional Chinese medicine in my country. Professors Chen Zhu and Wang Zhenyi used the drug to treat acute promyelocytic leukemia (APL), conquering APL for the first time, and clinical results showed that the patient's cure rate was as high as 95%. In addition to APL cells, ATO also has a certain killing effect on a variety of solid tumor cells, but its effect is far inferior to APL cells. Autophagy is one of the common responses of solid tumor cells against chemotherapy. Cells in a state of autophagy will reduce the activity of related enzymes needed to promote cell death, thereby greatly reducing the effect of chemotherapy. ATO can induce autophagy in a variety of sol...

Claims

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Application Information

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IPC IPC(8): A61K33/36A61K33/44A61K33/24A61K33/26A61P35/00
CPCA61K33/24A61K33/26A61K33/36A61K33/44A61K2300/00
Inventor 樊春海诸颖崔之芬
Owner SHANGHAI INST OF APPLIED PHYSICS - CHINESE ACAD OF SCI
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