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Application of berberine in preparing drug for overcoming drug resistance of chronic granulocytic leukemia or drug sensitizer for resisting chronic granulocytic leukemia

A technology for chronic myeloid cells and leukemia, which is applied in the field of biomedicine and can solve problems such as reducing the sensitivity of kinase inhibitors

Active Publication Date: 2016-06-01
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Mutations that alter the sequence of the BCR-ABL protein prevent binding of BCR-ABL kinase inhibitors, thereby reducing sensitivity to kinase inhibitors

Method used

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  • Application of berberine in preparing drug for overcoming drug resistance of chronic granulocytic leukemia or drug sensitizer for resisting chronic granulocytic leukemia
  • Application of berberine in preparing drug for overcoming drug resistance of chronic granulocytic leukemia or drug sensitizer for resisting chronic granulocytic leukemia
  • Application of berberine in preparing drug for overcoming drug resistance of chronic granulocytic leukemia or drug sensitizer for resisting chronic granulocytic leukemia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Berberine treatment of cell lines to detect cell viability

[0037] K562, SFO2 cells, BaF3-P210 and BaF3-P210-T315I cells were inoculated in RPMI-1640 medium containing 10% fetal bovine serum and antibiotics, and placed at 37°C, 5% CO 2 In the incubator, cultured under saturated humidity, and the medium was changed every other day; the K562, SFO2 cells, BaF3-P210 and BaF3-P210-T315I cells in the logarithmic growth phase were taken, and the cells in each group were divided into 1×10 8 The density of cell / L was inoculated in 96-well plate, 100 μL in each well; each cell was divided into berberine group, imatinib group, berberine+imatinib group and blank group, and each group was set 3 Add 100 μL of medium containing the corresponding concentration of drug to each well, and add blank medium to the control group, with a total volume of 200 μL; after adding the drug, put it in the incubator for 48h or 72h; cells were treated with the drug for 48h or 72h Then add 2...

Embodiment 2

[0043] Example 2 The effect of berberine on the colony formation of BaF3-P210 and BaF3-P210-T315I cells

[0044] In this example, a colony forming method using methylcellulose as a carrier was used to detect the self-renewal and proliferation abilities of cells. The experiment was divided into berberine group, imatinib group and blank group, the concentration of berberine was 5 μM, and the concentration of imatinib was 5 μM. Take the BaF3-P210 and BaF3-P210-T315I cells grown in the logarithmic phase, and press 1×10 3 cell / L (containing the corresponding concentration of treatment drugs) was inoculated in 2mmol / L of L-glutamine, containing 20% ​​fetal bovine serum by volume, 5 μmol / L β-mercaptoethanol and 0.9% methylcellulose in solid medium placed in a 37°C, 5% incubator. placed in the incubator for one week. One week later, the number of colonies was counted under an inverted microscope, and a colony contained at least 40 cells.

[0045] Figure 5 It is the analysis of c...

Embodiment 3

[0048] Embodiment 3RealtimePCR detection BCR-ABLmRNA level

[0049] (1) Extraction of total RNA: After treating BaF3-P210-T315I cells, BaF3-P210 cells and K562 cells for 24 hours with berberine, centrifuge at 1500 rpm for 5 minutes to collect the cells in a 1.5mLEP tube, add 500 μL Trizol reagent, blow and mix the cells evenly, Let stand on ice for 5 minutes; add 100 μL of chloroform (chloroform: Trizol=1:5), shake vigorously for 15 sec, and let stand on ice for 10 min; centrifuge at 12000 rpm at 4°C for 15 min, draw the supernatant into another EP tube, add an equal volume of isopropanol (about 200 μL), shake evenly, let stand at -20°C for 10min, centrifuge at 12000rpm at 4°C for 10min; discard the supernatant, and a white precipitate can be seen at the bottom of the EP tube; add 500μL of ethanol with a volume fraction of 75% to each tube (use RNase-freeddH 2 O preparation), wash the RNA precipitation; after washing, centrifuge at 12000rpm, 4°C for 5min, repeat this step 3 t...

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Abstract

The invention belongs to the field of biological medicine, and particularly relates to application of berberine in preparing a drug for overcoming the drug resistance of chronic granulocytic leukemia or a drug sensitizer for resisting chronic granulocytic leukemia. Berberine can improve the sensitivity of chronic granulocytic leukemia cells to imatinib and overcome the multi-drug resistance of chronic granulocytic leukemia cells to imatinib. Berberine degrades BCR-ABL protein mainly through a ubiquitination mechanism, and thus the multi-drug resistance of chronic granulocytic leukemia caused by T315I mutation is overcome.

Description

technical field [0001] The invention belongs to the field of biomedicine, and specifically relates to the application of berberine in the preparation of drugs for overcoming chronic myeloid leukemia drug resistance or anti-chronic myeloid leukemia drug sensitizers. Background technique [0002] Chronic Myeloid Leukemia (CML) is a myeloid malignant clonal disease characterized by abnormal transformation of hematopoietic stem cells. Its typical genetic characteristic is t(9;22)(q34;q11) translocation, which produces BCR / ABL fusion gene. Globally, CML accounts for 15-20% of all adult leukemias, and its incidence ranks third among leukemias in my country. [0003] Over the past 20 years, the disease progression and mortality rates of CML have remained high. In recent years, with the advent of the molecular targeted therapy drug tyrosine kinase inhibitor (TKI) imatinib, the treatment of CML has undergone revolutionary progress, the 5-year event-free survival (EFS) and overall s...

Claims

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Application Information

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IPC IPC(8): A61K31/4375A61P35/02A61K36/718A61K36/756A61K36/29
CPCA61K31/4375A61K36/29A61K36/718A61K36/756
Inventor 费嘉丰茂晓阴钊古春明杨菊华
Owner JINAN UNIVERSITY
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