Medical applications of four buxus alkaloids compounds

A technology of alkaloids and compounds, applied in the field of medicine, can solve problems such as slow heartbeat

Inactive Publication Date: 2013-06-12
KUNMING UNIV OF SCI & TECH +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The total plant alkaloids of this genus generally have the effect of lowering blood pressure; the boxwood alkaloids act on the vagus nerve and the heart, resulting in slow heartbeat, local ischemia, sinoventricular block and myocardial injury; some plant total alkaloids also have an effect on cholinesterase Inhibitory effect, there is no report about the use of boxwood alkaloids in the treatment of tumors

Method used

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  • Medical applications of four buxus alkaloids compounds
  • Medical applications of four buxus alkaloids compounds
  • Medical applications of four buxus alkaloids compounds

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: Separation, extraction and identification of four compounds

[0031] Euonymus microphylla collected from Kunming Botanical Garden ( Buxus. microphylla ) aboveground part 14Kg, the plant species name was identified by researcher Qiu Minghua from Kunming Institute of Botany, and the crude drug specimens are stored in the State Key Laboratory of Phytochemistry and Western Plant Resources Sustainable Utilization, Kunming Institute of Botany, Chinese Academy of Sciences.

[0032] 14Kg of aerial parts of Euonymus microphylla, crushed and extracted with 70% acetone at room temperature for 3 times, each time for 2 days; combined extracts, concentrated under reduced pressure until no acetone was evaporated to obtain 800 g of extract, diluted with 3000ml of acidic water to adjust pH =2, extracted 3 times with 3000ml ethyl acetate respectively to obtain the non-alkaloid part; alkalized the acidic aqueous solution to PH=10, extracted 3 times with 3000ml chloroform to ...

Embodiment 2

[0040] Embodiment 2: Antitumor activity experiment of the compound of the present invention

[0041] 1. Construction of screening cells: wild-type mice were used as carriers, and p53 gene-deleted mouse embryonic fibroblasts (p53 - / - cells) to construct mouse tumor cells (p53 - / - +S+Ras, p53 - / - +V+Ras), the specific steps are as follows:

[0042] (1) Construction of vectors for Ras mutant genes and vectors expressing p53S mutant genes

[0043] For the construction method of the vector, see the method in patent ZL200910095184.3, that is, the p53S or Ras mutant gene cDNA is connected into the PQCXIP vector by conventional genetic engineering methods, and the p53S or Ras sequence constructed in the vector is verified by sequencing. The vector used in p53 - / - The p53S or Ras mutant gene is introduced into the cells.

[0044] (2) Transfection of p53S or Ras mutant gene vector

[0045] (a) Preparation of phoenix cells

[0046] The day before transfection, phoenix cells (h...

Embodiment 4

[0062] Example 4: Western blot experiment

[0063] 1. Protein sample pretreatment and concentration determination

[0064] (1) The p53 in the logarithmic growth phase - / - +V+Ras, p53 - / -+S+Ras cells were treated with 5 μM KBR18, KBA01, KBA02 and control DMSO for 8h, and after 16h and 24h, the cells were gently scraped off the plate, and put into a 15mL centrifuge tube together with the medium, at 4°C, 4000 rpm ( Centrifuge at 3082′g for 5min, discard the supernatant, suspend the cell pellet gently with 1mL 1×PBS, then transfer the cell suspension into a 1.5mL centrifuge tube, centrifuge at 13000 rpm (12470′g) for 1min, and aspirate the supernatant Clean, the cell pellets are stored at -80°C for later use;

[0065] (2) Lyse the cell pellet with 0.2mL cell lysate (the volume of the lysate depends on the amount of the cell pellet, generally 3-5 times the cell volume is better, if too much, the concentration of the cell lysate is too dilute, and the sample volume If too litt...

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Abstract

The invention discloses medical applications of four buxus alkaloids compounds in the preparation of antitumor medicines. Experiment results show that the four compounds KBA01, KBA02, KBA03 and KBR18, under the condition of concentration without toxicity to wild normal mouse embryonic fibroblast cells, can selectively kill mouse tumor cells with tumor specific mutant genes p53 and Ras; accordingly, the compounds are very strong in killing activity to human colon cancer cell strains with p53 mutants, but not strong in killing activity to human wild colon cancer cell strains with p53 mutants; and as shown in a GFP (Green Fluorescence Protein) reporter vector screening system of a gene promoter, the compounds KBA02 and KBA03 can activate a promoter of a cancer suppressor gene p16. The characteristics indicate that the compounds from the four buxus plant origins have application potentials for preparing personalized treatment medicines aiming at the tumor mutant genes p53 and Ras as well as the cancer suppressor gene p16.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to the application of four kinds of boxwood alkaloid compounds in the preparation of antitumor drugs. Background technique [0002] Tumor is a kind of fatal disease that seriously threatens human health. Its pathogenesis has not been fully elucidated, and the treatment effect is not satisfactory. Most of the drugs currently routinely used in tumor therapy are designed for the regulation of DNA replication or cytoskeleton in the process of cell proliferation. While these drugs kill rapidly proliferating tumor cells, they are also highly toxic to normal proliferating cells (such as bone marrow stem cells, etc.) On the contrary, the resistance of the tumor decreases, and it is difficult to obtain a good therapeutic effect. Therefore, using molecular targeted screening technology, targeting tumor cell-specific gene mutation products, screening small molecular compounds that can inh...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/57A61K31/56A61P35/00
Inventor 罗瑛邱明华张继虹阎玉鑫苏红陆思千贾舒婷吴晓明
Owner KUNMING UNIV OF SCI & TECH
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