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Drug target prediction and drug comprehensive evaluation method

An evaluation method and drug technology, applied in biochemical equipment and methods, drug combinations, pharmaceutical formulations, etc., can solve the problem that the coverage of evaluation indicators is difficult to achieve ideal results.

Active Publication Date: 2017-08-22
北京极客基因科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Although the high-content system can evaluate potential drug candidates from multiple dimensions, the coverage of its evaluation indicators is still difficult to achieve the desired effect

Method used

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  • Drug target prediction and drug comprehensive evaluation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0075] Example 1. Construction of a multi-label whole transcriptome sequencing library

[0076] (1) Cell culture

[0077] 1. Culture and passage of HepG2 liver cancer cell line

[0078] (1) The HepG2 liver cancer cell line was cultured in DMEM-HG medium containing 10% FBS to a density close to 80%.

[0079] (2) Wash twice with PBS buffer to remove dead cells on the upper layer, add an appropriate amount of Trypsin to digest at 37°C for 3 minutes, then neutralize with DMEM-HG medium with 10% FBS, centrifuge and remove the supernatant.

example 1

[0080] (3) Resuspend in DMEM-HG medium with 10% FBS, count and pass, the passage ratio is 1:6.

[0081] 2. Test the effectiveness of FDA-approved targeted drugs on the HepG2 liver cancer cell line

[0082] (1) The HepG2 liver cancer cell line was cultured in DMEM-HG medium containing 10% FBS to a density close to 80%.

[0083] (2) Wash twice with PBS buffer to remove dead cells on the upper layer, add an appropriate amount of Trypsin to digest at 37°C for 3 minutes, then neutralize with DMEM-HG medium with 10% FBS, centrifuge and remove the supernatant.

[0084] (3) Resuspend in DMEM-HG medium with 10% FBS, count and passage, passage density: 1.0x10^5 / well.

[0085] (4) Preparation of the drug to be tested: Dilute the drug to an appropriate concentration, usually 10 mM. The test concentration is set to 1000x, 5000x, 10000x.

[0086] (5) After passage for 24 hours, add the drug to be tested when the confluence of the cells reaches about 60%. Three parallel wells were set up...

Embodiment 2

[0141] Example 2. Sequencing and result analysis of multi-label whole transcriptome sequencing library

[0142] (1) The library constructed in Example 1 was subjected to paired-end sequencing on the Illumina sequencing platform.

[0143] (2) Analyze the above sequencing results:

[0144] 1. Cluster analysis of sequencing results

[0145] Figure 5 It is the clustering analysis of the HepG2 gene expression of the drug to be tested for 72 hours. It can be seen that the targeted drug drug #2R428 (the target is Axl) and #5Crizotinib (the target is c-Met) have a lethal effect on liver cancer cell HepG2. similar expressive features. Unlike #7 Sunitinib (targeting RTK) and #3 Sorafenib (targeting ERK), a drug known to target liver cancer cells, it shows that their mechanisms of killing liver cancer cells HepG2 are different.

[0146] 2. Spantree analysis of sequencing results

[0147] Figure 6 It is the Spantree analysis of HepG2 gene expression of the drug to be tested for 72...

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Abstract

The invention provides a drug target prediction or drug evaluation method, a multi-label whole transcriptome sequencing library construction method and a multi-label whole transcriptome sequencing library constructed by the method. The method and the library of the invention are suitable for single cells or a few cells, are low-cost, have low quantity demand on samples, and can help realize high throughput accurate evaluation of the action of a drug in a living system. The invention relates to a new generation high-accuracy and high-dimensionality drug screening and evaluation system capable of comprehensive systematic research on the mechanism of drug action and can predict potential combination effect among different drugs and possible new drug research and development target so as to guide new drug research and development and clinical administration.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for drug discovery and evaluation, target prediction and medication guidance based on a single-cell or small-number-of-cell transcriptome sequencing method. Background technique [0002] At present, the commonly used drug screening and evaluation systems in the world are mainly divided into high-throughput screening and high-content screening. Among them, the classic high-throughput screening mainly uses robots and computers to realize fully automated biological and pharmacological experiments in the pharmaceutical and biological industries, so as to quickly evaluate a large number of unknown compounds to discover potential candidate drugs with biological activity. [0003] However, due to the limitations of high-throughput screening technology itself, it is very dependent on a highly sensitive single reporter system. Although high-throughput screening can comple...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C40B50/06C40B40/06A61K31/404A61K31/502A61K31/44A61K31/4545A61P35/00
CPCA61K31/404A61K31/44A61K31/4545A61K31/502C12Q1/6806C12Q1/6869C40B40/06C40B50/06
Inventor 罗昊澍包仁艳
Owner 北京极客基因科技有限公司
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