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A kit and application for screening drug developmental toxicity based on c/ebpα and igf1r genes

A kit and gene technology, applied in the field of drug toxicology, can solve the problems of unclear developmental toxicity of drugs, lack of theoretical system of occurrence mechanism, etc., and achieve the effects of high sensitivity, fast detection process and wide application range.

Active Publication Date: 2020-12-01
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In summary, the in vitro evaluation system of drug developmental toxicity has great limitations, which are mainly related to the unclear performance of drug developmental toxicity and the lack of a theoretical system for the mechanism of occurrence. Therefore, a high-throughput drug developmental toxicity in vitro evaluation system based on toxicity mechanisms and molecular targets remains to be studied.

Method used

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  • A kit and application for screening drug developmental toxicity based on c/ebpα and igf1r genes
  • A kit and application for screening drug developmental toxicity based on c/ebpα and igf1r genes
  • A kit and application for screening drug developmental toxicity based on c/ebpα and igf1r genes

Examples

Experimental program
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Effect test

Embodiment 1

[0050] The effect of a kit for detecting drug developmental toxicity based on C / EBPa and IGF1R genes on the developmental toxicity drug methimazole in the L02 cell line model.

[0051] The kit consists of detecting intracellular C / EBPa and IGF1R gene expression, and simultaneously detecting the luciferase activity of the IGFIR promoter and luciferase reporter gene. The above-mentioned kit for detecting drug developmental toxicity based on C / EBPa and IGF1R genes is applied to the developmental toxicity drug methimazole in the L02 cell line model as follows:

[0052] 1 Preparation of human fetal L02 cells

[0053] 1.1 Take out the cell cryopreservation tube from the liquid nitrogen tank, put it into a 37°C water bath quickly, and shake it from time to time to thaw as soon as possible.

[0054]1.2 After complete thawing, centrifuge at 1300rpm for 3min, wipe and disinfect the cryopreservation tube with 75% alcohol, and then move it to a biological safety cabinet.

[0055] 1.3 As...

Embodiment 2

[0094] The effect of a kit for detecting drug developmental toxicity based on C / EBPa and IGF1R genes on the developmental toxicity drug methimazole in WJ-MSCs hepatic-like differentiation cell model.

[0095] 1. WJ-MSCs primary extraction and hepatic differentiation

[0096] 1.1 WJ-MSCs primary extraction

[0097] Under aseptic conditions in the operating room, the umbilical cord of the fetus delivered by cesarean section was taken out, the blood was washed, soaked in pre-cooled saline, and brought back to the laboratory. The umbilical cord was taken out in the ultra-clean workbench, and all separation processes were performed in pre-cooled saline. Cut into 4-6cm / section, and wash away the blood with a syringe. Remove the umbilical cord adventitia, umbilical artery, and umbilical vein, and peel off Wharton's glue: first, fix the two ends of the umbilical cord on a wooden board with tacks, and use ophthalmic scissors to separate the adventitia longitudinally from one end, and...

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Abstract

The invention relates to a kit for screening drug developmental toxicity based on a C / EBP[alpha]-IGF1R gene and an application thereof. The kit contains: primers for detecting expression of C / EBP[alpha] and IGF1R gene mRNA; drug acting object mammalian cell lines or differentiated cells, and cell lines or differentiated cells having a plasmid containing an IGF1R promoter and a luciferase reportergene transferred into cells; a cell lysate; and luciferase. The determination standard of the kit comprises improvement of intracellular C / EBP[alpha] gene expression of a to-be-screened compound, reduction of IGF1R gene expression, and reduction of the activity of the luciferase reporter gene in the cells containing the IGF1R promoter-reporter gene, and the to-be-screened compound is suggested tohave developmental toxicity. The kit is novel, efficient and reliable, and can be used for high-throughput drug screening, and is of great significance for rapid detection of drugs and other compoundswith developmental toxicity.

Description

technical field [0001] The invention belongs to the field of drug toxicology, and relates to a kit and application for screening developmental toxicity of drugs or other compounds based on C / EBPa and IGF1R genes. Background technique [0002] Drug developmental toxicity refers to the structural or functional damage in offspring before sexual maturity caused by drug factors, including structural deformity, dysfunction, growth retardation and even death. Drug developmental toxicity is one of the important contents of drug safety evaluation. At present, internationally recognized guidelines related to developmental toxicity of drugs include "Organization for Economic Co-operation and Development (OECD) Chemical Substance Testing Guideline 414-Developmental Toxicity Studies During Pregnancy" and "Guidelines for Drug Registration (ICH)". The results of reproductive experiments are used as the main indicator to evaluate the developmental toxicity of drugs, but there are disadvant...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883C12Q1/66
CPCC12Q1/66C12Q1/6883C12Q2600/142C12Q2600/158
Inventor 汪晖文印宪齐勇建何波王桂花
Owner WUHAN UNIV
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