Application of c-Abl kinase inhibitor in FoxM1 high-expression tumor treatment

A kinase inhibitor, 1. The technology of c-abl, which is applied in the field of biomedicine, can solve the problems of unclear inhibitory effect on solid tumors, achieve the effect of inhibiting tumor formation, expanding clinical indications, and broad application prospects

Inactive Publication Date: 2019-08-09
ACADEMY OF MILITARY MEDICAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Nilotinib is mostly used in the treatment of blood cancers such as leukemia in clinical practice, and its inhibitory effect on solid tumors is not yet clear

Method used

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  • Application of c-Abl kinase inhibitor in FoxM1 high-expression tumor treatment
  • Application of c-Abl kinase inhibitor in FoxM1 high-expression tumor treatment
  • Application of c-Abl kinase inhibitor in FoxM1 high-expression tumor treatment

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1, Construction of Control Hela-S3 and Hela-FoxM1Y575F cell lines and inoculation of nude mice

[0033] 1. Construction of Control Hela-S3 and Hela-FoxM1Y575F cell lines

[0034] The Hela-FoxM1Y575F cell line is a homozygous cell line in which the 575th tyrosine residue of FoxM1 is mutated to phenylalanine.

[0035] 1. Construction of pSpCas9-FoxM1Y575F expression plasmid

[0036] Centering on the codon encoding the 575th tyrosine of FoxM1, intercept the FoxM1 gene sequence of 100 bp upstream and downstream, a total of 203 bp, and submit it to the CRISPR online design tool (http: / / crispr.mit.edu / ) to design and generate the sgRNA sequence 5'-CTCCCAGCTCAGCTACTCCC-3', and CACCG is added to the 5' end of this sequence, and AAAC is added to the 5' end of the antisense strand. The DNA was mixed with the pSpCas9(BB)-2A-Puro plasmid digested by BbsⅠ, ligated under the action of T4DNA ligase, and the ligated product was transformed into DH5α competent cells, and the p...

Embodiment 2

[0051] Embodiment 2, nude mice Nilotinib intragastric administration

[0052] Administration began on the 2nd day after nude mice were inoculated with cells. a Dissolve nilotinib in dimethyl sulfoxide at a concentration of 15 μg / μl, and then dissolve this solution in physiological saline at a volume ratio of 1:1.67 to prepare a solution of 5.6 μg / μl. b The nude mice administered with nilotinib were intragastrically administered with a volume of 200 μl per day using a syringe and a gavage needle. c The solution given to the control group was physiological saline containing 37% (volume ratio) dimethyl sulfoxide, 200 μl per day for each mouse. Nude mice in each group were administered continuously for 21 days. 8 nude mice in each group.

Embodiment 3

[0053] Example 3, Detection of Tumor Cell Colony Formation Ability

[0054] Cell colony formation assay is often used to detect the division and proliferation ability of tumor cells. Use trypsin to digest ControlHela-S3 cells and Hela-FoxM1Y575F cells. After the digestion is complete, add DMEM medium to neutralize the trypsin, and use a pipette to repeatedly pipette to form a single-cell suspension. Use a cell counter to detect the cell concentration, according to 1 × 10 3 The inoculum amount of cells / well was inoculated in a six-well plate and placed in 37°C, 5% CO 2 Culture in an incubator, and replace with fresh medium every 2-3 days. The cells were cultured for about 10 days to form a macroscopic colony. Remove the medium and add 4% paraformaldehyde to fix the cells for about 20 minutes. After removing the 4% paraformaldehyde, wash the cells twice with 1×PBS buffer. Incubate staining solution at room temperature for 30 minutes, then wash with light buffered deionized wa...

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Abstract

The invention discloses application of a c-Abl kinase inhibitor in FoxM1 high-expression tumor treatment and provides application of the c-Abl kinase inhibitor, especially nilotinib, in preparing products for FoxM1 high-expression tumor treatment. It is proved that nilotinib can remarkably inhibit growth of cervical cancer cells, and after FoxM1 phosphorylation sites which c-Abl depend on are mutated, the tumor formation capability of the cervical cancer cells can be remarkably inhibited. The potential of the c-Abl kinase inhibitor, namely nilotinib, in FoxM1 high-expression solid tumor treatment is explored, a new theoretical support and experimental basis are provided for clinical application of nilotinib, clinical adaptability of nilotinib is expected to be expanded, and nilotinib has wide application prospects in the field of tumor treatment.

Description

technical field [0001] The invention relates to the field of biomedicine, and relates to the application of c-Abl kinase inhibitor in the treatment of tumors with high FoxM1 expression. Background technique [0002] c-Abl non-receptor tyrosine kinase can activate related signaling pathways by phosphorylating substrate proteins. Under normal physiological conditions, its kinase activity is in a state of autoinhibition, and the activated c-Abl kinase is more likely to be degraded through the ubiquitin-proteasome pathway. In chronic myeloid leukemia, the Piladelphia chromosome formed due to chromosomal heterolocation leads to the fusion of ABL and BCR genes, and the formed BCR-ABL has continuously activated kinase activity. Nilotinib is a second-generation tyrosine kinase inhibitor that inhibits the kinase activity of c-Abl and prevents the phosphorylation of c-Abl substrates by competitively binding to the ATP-binding site on BCR-Abl kinase , thereby inhibiting related signa...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/506A61P35/00
CPCA61K31/506A61P35/00
Inventor 刘萱袁永倩董钦才曹诚靳彦文张部昌
Owner ACADEMY OF MILITARY MEDICAL SCI
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