Tumor targeted polypeptide-adriamycin amycin derivative as well as preparation method and application thereof

A technology of tumor targeting and doxorubicin, which is applied in the direction of antineoplastic drugs, drug combinations, pharmaceutical formulations, etc., can solve the problems of high transferrin concentration and achieve increased drug concentration, small steric hindrance, high purity and high yield Effect

Active Publication Date: 2014-07-09
南京明捷生物医药检测有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the body's endogenous transferrin concentration is high, and competition inhibits the targeted transport of transferrin-modified drug delivery systems.

Method used

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  • Tumor targeted polypeptide-adriamycin amycin derivative as well as preparation method and application thereof
  • Tumor targeted polypeptide-adriamycin amycin derivative as well as preparation method and application thereof
  • Tumor targeted polypeptide-adriamycin amycin derivative as well as preparation method and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Synthesis and purification of embodiment 1 polypeptide-doxorubicin complex

[0036] (1) Preparation of T10-DOXO-EMCH and T15-DOXO-EMCH

[0037] Weigh 0.8 mg of T10 peptide, dissolve it in 600 μL of pH7.8-8.2 phosphate buffer, then weigh 1.13 mg of DOXO-EMCH, and dissolve it in 600 μL of N,N'-dimethylformamide aqueous solution, where N , the volume ratio of N'-dimethylformamide to water is 1:1, drop into the polypeptide solution at a rate of 25 μL / min, react at room temperature for 1.5 hours, and generate T10-DOXO-EMCH, transfer the solution to a centrifuge tube, Centrifuge at 8000×g for 5 min to remove unreacted substances to obtain a crude product.

[0038] The structure of doxorubicin prodrug (6-maleimidocaproyl) hydrazone derivative—DOXO-EMCH is as follows:

[0039]

[0040]Weigh 1.23mg of T15 peptide, dissolve in 600μL pH7.8~8.2 phosphate buffer, then weigh 1.13mg of DOXO-EMCH, dissolve in 600μL N,N'-dimethylformamide aqueous solution, where N , the volume rat...

Embodiment 2

[0051] Example 2 Intracellular Adriamycin Accumulation Experiment

[0052] MCF-7 / ADR cells at 1×10 5 The density per well was inoculated in a 6-well culture plate, and after incubation at 37°C and 5% carbon dioxide for 24 hours, DOX and its derivatives T10-DOXO-EMCH and T15-DOXO-EMCH were added with a final concentration of 1.5 μM. The effect is 12h. Discard the drug-containing culture medium, wash the cells 3 times with cold PBS, and observe the treated cells in each group under a fluorescent microscope. The results are shown in figure 2 . After MCF-7 / ADR cells were treated with different drugs, the fluorescence intensity in the T10-DOXO-EMCH and T15-DOXO-EMCH groups was significantly higher than that in the DOX group.

Embodiment 3

[0053] Embodiment 3MTT method measures cell viability test

[0054] Digest MCF-7 / ADR cells in good condition and dilute to 2×10 with culture medium 4 cells / mL cell density, after blowing evenly, add 200 μL of cell suspension to each well of a 96-well plate, and incubate at 37°C and 5% carbon dioxide for 24 hours to make it adhere to the wall. Replace with 200 μL DOX, T10-DOXO-EMCH and T15-DOXO-EMCH drug-containing culture medium, set 5 parallel wells for each concentration, and set a blank control group at the same time, place them in an incubator for 72 hours, and then add MTT solution (5 mg / mL) was 20 μL, continued to incubate for 4 h, the culture medium was sucked off, DMSO 200 μL was added, and the microplate shaker was shaken for 10 min at room temperature, and the absorbance value of each well was measured at 490 nm in an enzyme-linked immunosorbent assay instrument. Taking the control cells as 100%, the survival rate of the cells in the administration group relative to...

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Abstract

The invention relates to a tumor targeted polypeptide-adriamycin amycin derivative as well as a preparation method and application thereof. The compound is prepared by virtue of the following steps: dissolving polypeptide in a phosphate buffer solution with the pH value of 7.8-8.2; then, dissolving DOXO-EMCH in a solvent to obtain a solution, wherein the polypeptide is T10 peptide or T15 peptide and the solvent is a mixture of water and N,N'-dimethylformamide; dropwise adding the DOXO-EMCH solution into a polypeptide solution; stirring and reacting away from sunlight at room temperature; then, centrifugalizing the solution to remove precipitate to obtain a coarse product; separating and purifying the coarse product by preparative chromatography, and centrifugalizing and concentrating to obtain a fine product T10-DOX-EMCH or T15-DOX-EMCH. According to the adriamycin amycin hydrazone compound provided by the invention, the hydrazone bond is selectively broken in an acidic microenvironment of tumor to release adriamycin amycin with pharmacological activity, so that the curative effect of medicine is improved and the drug tolerance of tumor is reversed.

Description

technical field [0001] The invention belongs to the technical field of biology and medicine, and in particular relates to a polypeptide with anticancer activity-doxorubicin derivatives and its preparation method and application. Background technique [0002] Doxorubicin (DOX), an anthracycline compound, is one of the most effective anticancer drugs and is widely used clinically to treat acute leukemia, breast cancer, lymphoma and various solid tumors. But like other anticancer drugs, the clinical application of doxorubicin is severely limited due to its easy drug resistance and poor selectivity. In the past 20 years, a large number of structural modifications and modifications have been carried out for doxorubicin in order to enhance tumor targeting and improve efficacy. [0003] The drug resistance and low selectivity of anticancer drugs can be obtained by using various polypeptides or proteins as carriers, using the receptor-mediated endocytosis mechanism of the carrier, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K47/48A61K31/704A61P35/00A61K47/64A61K47/66
Inventor 陈芸徐金慧盛媛许飞飞方丹君尤一雯
Owner 南京明捷生物医药检测有限公司
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