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Rapid high-throughput screening model for antiviral drugs

A high-throughput, drug technology, applied in the direction of antiviral agents, pharmaceutical formulations, and the determination/inspection of microorganisms. Effective screening rate, simplification of screening procedures, and the effect of improving screening rate

Active Publication Date: 2011-03-09
HUNAN DANWEI BILOGICALTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the animal screening method also has disadvantages: when using animal models for drug screening, due to the particularity of animals, the drug screening process mainly depends on manual operations, and only limited samples can be screened, so that the overall animal model screening of new drugs has the advantages Obvious limitations, low efficiency, high cost, long time required for animal experiments, high labor intensity, high technical requirements for operation, large amount of test samples (about 5g)
The pharmacological effect of the drug is exerted through the drug target of the body cells or pathogens. Whether the drug for internal use is administered through any route of administration, such as oral administration or injection, it needs to go through the process of absorption, metabolism, distribution and excretion. Affect the effect of drugs, such as metabolism affects the molecular structure and effect of drugs through oxidation, decomposition, and combination. Enhancement, such as the metabolism of the inactive anticancer drug azathioprine to the pharmacologically active 6-thiopurine
Conventional in vitro cell or pathogen culture drug screening models, because the drug does not go through in vivo absorption, distribution, metabolism and other processes, but directly acts on cells or pathogens; therefore, it cannot truly reflect the role of the drug, and there is uncertainty and non-correspondence. Some drugs or drug ingredients that are effective in vitro experiments are not effective in clinical experiments
Usually, there is a big deviation between the results of in vitro drug screening and the clinical effect; at the same time, the conventional in vitro cell culture drug screening model needs to measure the safe dosage of cultured cells for each drug, which has the disadvantages of heavy workload, complicated operation and high cost.

Method used

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  • Rapid high-throughput screening model for antiviral drugs
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  • Rapid high-throughput screening model for antiviral drugs

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Determination of Toxic Effects of Chinese Herbal Medicine Extracts on Cultured Cells

[0026] In order to understand and compare the toxic effects of direct drug addition on cells by conventional methods, the toxic effects and safe concentrations of water extracts of 21 Chinese herbal medicines used in the test on Marc-145 cells were measured. The method is to roughly extract 21 kinds of Chinese herbal medicines with water respectively. The water extracts of each Chinese herbal medicine are sterilized and diluted with cell maintenance solution, and added to 96-well culture plates full of cells, with 8 wells per concentration and 100ul per well. 37°C 5% CO 2 Cultivate in the incubator for 72h, observe CPE every day. The results show that various extracts have different degrees of toxic effects on the cells, and the effects increase with the increase of the concentration, and the side effects of different Chinese herbal medicines are different. Add 0.625-1.25% (V / V) or m...

Embodiment 2

[0028] Determination of Safe Concentration of Mouse Serum on Marc145 Cells

[0029] Mouse serum was inactivated at 56°C for 30 minutes, diluted to 80% with maintenance solution, sterilized by filtration through a 0.22 μm microporous membrane, and diluted to different concentrations with maintenance solution. The dilution concentrations of mouse serum were 80%, 40%, 20%, 10%, 5%, 2.5%, 1.25%, 0.625%, 0.312%, from high concentration to low concentration were added to well-grown monolayer cells, 100 μl per well, repeated for each concentration In 8 wells, a cell control group was set, and they were cultured in a 37°C incubator for 72 hours. The CPE was observed with an inverted microscope every day and the results were recorded. The final recorded results were regarded as the final results. At the same time, the OD value measured by MTT method was used to calculate the cell viability. The specific operation is as follows: after 72 hours of treatment, suck out the residual mouse ...

Embodiment 3

[0032] Protection of mouse serum containing Chinese herbal medicine against PRRSV virus infection in Marc-145 cells and screening of antiviral drugs

[0033] 1 test material

[0034] Marc-145 cells (African rhesus monkey kidney cells): donated by the School of Veterinary Medicine, Hunan Agricultural University;

[0035] American porcine reproductive-respiratory syndrome virus (PRRSV): donated by the School of Veterinary Medicine, Hunan Agricultural University;

[0036] Chinese herbal medicine: 21 kinds of Chinese herbal medicine (Guanzhong, Zhonglou, betel nut, star anise lotus, Eucommia, Radix Isatidis, Phellodendron, Aracea, Pinellia, Folium Isatidis, Zihuadiding, Xuanhu, Scutellaria barbata, Cornus officinalis, Forsythia, Cyanorrhizae, oregano, verbena, magma fruit, cnidium, nuxychia, etc.) were purchased from Hunan Yangtianhe Pharmacy, extracted three times by traditional decoction, combined and concentrated, filtered and stored at 4°C;

[0037] 2 test method

[0038] T...

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Abstract

A establishment method of a novel high-throughput screening model is provided in the invention, which aims to solve the problems of nondeterminacy and incorrespondence exsiting in general in vitro high-throughput screening method. The model provided by the invention furthest simulates animal physiological environment, thus the accuracy of drug screening is increased and the results obtained by the method of in-vitro screening are closer to that of clinical application. The technical scheme of the present invention is as follows: Animal medicated serum is obtained by absorpting and separating drug active ingredients during the process of in vivo circulation of animals, thereby the medicated serum is added to cell culture media or maintenance media after drug is subject to the process of in vivo metabolism of animals. Cell activity is calculated by utilizing micro cell culture method, cytopathic effects (CPE) measurement and MTT method.

Description

technical field [0001] The invention relates to the field of drug screening models, in particular to a method for establishing a rapid high-throughput screening model of antiviral drugs, which can be applied to drug screening. Background technique [0002] Drug screening model (drug screening assay method) is an experimental method used to prove that a certain substance has pharmacological activity (biological activity, therapeutic effect), and these experimental methods are one of the important conditions for finding and discovering drugs. In the long-term practice of searching for drugs, people have established a large number of various models for new drug screening, which can be roughly divided into classic animal experiment methods, molecular biology screening methods, high-throughput drug screening methods, and high-content drug screening techniques. [0003] The so-called animal screening method is to use animals as the observation objects of drug screening, to prove t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K49/00C12Q1/02A61P31/12
Inventor 肖兵南
Owner HUNAN DANWEI BILOGICALTECH
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