Rapid screening method for galactosyl ceramide transferase inhibitor drug
A technology of galactosylceramide and screening method, which is applied to measurement devices, material analysis by electromagnetic means, instruments, etc., can solve the problems of complex effect evaluation and slow screening, and achieve the effect of fast and stable screening method
Inactive Publication Date: 2019-05-03
辉源生物科技(上海)有限公司
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Problems solved by technology
However, the effect evaluation of existing ceramide galactosyltransferase inhibitor drugs is relatively complicated, which makes the screening of highly effective ceramide galactosyltransferase inhibitor drugs very slow. Therefore, galactosylceramide transferase (CGT) inhibitory Therefore, rapid screening methods for agents are particularly important
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Embodiment 1
[0024] A rapid screening method for galactosylceramide transferase inhibitor drugs, comprising the following steps:
[0025] (1) Construction of human galactosylceramide transferase overexpression vector
[0026] The ribonucleic acid of human galactosylceramide transferase that can be expressed through artificial synthesis was constructed on the mammalian cell expression vector pDNA3.1(+) (ThermoFisher Scientific, Grand Island, NY)
[0027] (2) Using non-human cells for cell transfection of human galactosylceramide transferase overexpression vector and preparation of cell lysate protein
[0028] The human galactosylceramide transferase overexpression vector was transfected into non-human cells (Chinese vole ovary cells; CHO). After transfection, cells were lysed using Mammalian Protein Extraction Reagent (M-PER) and cell lysate proteins were extracted;
[0029] 3. Development of an in vitro drug screening method using cell lysates containing human galactosylceramide transfer...
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The invention relates to a rapid screening method for a galactosyl ceramide transferase inhibitor drug. The method comprises: step one, artificially synthesizing human galactosyl ceramide transferaseribonucleic acid and then constructing ribonucleic acid of the alactosyl ceramide transferase on a mammalian cell expression vector pDNA3.1(+) to obtain a human galactosyl ceramide transferase overexpression vector; step two, carrying out cell transfection of the human galactosyl ceramide transferase overexpression vector on non-human cells, carrying out cell lysis, and extracting cell lysis protein; and step three, placing the extracted cell lysis protein and a reaction substrate in a reaction buffer solution, adding a to-be-screened galactosyl ceramide transferase inhibitor drug, carrying out reaction, detecting contents of a substrate and a reaction product, and determining the inhibitory effect of the to-be-screened galactosyl ceramide transferase inhibitor drug. Compared to the priorart, the rapid screening method has the following advantage: establishing a rapid and stable screening mode.
Description
technical field [0001] The invention relates to the technical field of drug screening, in particular to a rapid screening method for galactose ceramide transferase inhibitor drugs. Background technique [0002] Krabbe disease, also known as Krabbe disease, was first reported by Danish pediatrician Krabbe in 1916. According to its clinical features, it is also called infantile familial diffuse sclerosis. Krabbe disease is an autosomal recessive genetic metabolic disease, and the mutation gene is located at 14p. The gene defect of Krabbe disease causes galactocerebroside-β-galactosidase deficiency, which is a genetic metabolic disease that leads to the deposition of many galactocerebrosides in the white matter of the brain. The prognosis of this disease is extremely poor, and infantile patients often die within 1 year of age. Those with late onset can survive until about 10 years old. Since galactose ceramide transferase can convert ceramide into galactose cerebroside, inh...
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IPC IPC(8): G01N27/62
Inventor 白艳秋吴治昊王朝弘刘豫屏
Owner 辉源生物科技(上海)有限公司