Artificially synthesized cationic peptide and applications thereof in preparing anti-tumor drug

An anti-tumor drug, artificial synthesis technology, applied in the biological field, can solve the problems of large molecular weight, poor tissue penetration, low anti-tumor activity, etc., to achieve a small molecular weight, high anti-tumor activity, easy to penetrate tumor tissue Effect

Active Publication Date: 2014-08-20
HARBIN MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0013] In order to overcome the shortcomings of existing polypeptide anticancer drugs such as low antitumor activity, large molecular weight, and poor tissue penetration, the inventors of the present inven...

Method used

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  • Artificially synthesized cationic peptide and applications thereof in preparing anti-tumor drug
  • Artificially synthesized cationic peptide and applications thereof in preparing anti-tumor drug
  • Artificially synthesized cationic peptide and applications thereof in preparing anti-tumor drug

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Embodiment 1 Artificial chemical synthesis method prepares AIK on a large scale

[0028] 1. Chemical synthesis of AIK:

[0029]Synthesized by Beijing Saibaisheng Gene Technology Co., Ltd. Using the ABI433peptide synthesizer full-brake peptide synthesizer from Applied Biosystems, the solid-phase peptide synthesis method was used to continuously add, react, and synthesize from the C-terminus-carboxyl-terminus to the N-terminus-amino-terminus according to the sequence of AIK, and finally synthesized 100mg25 peptide AIK. The amino acid sequence is shown in SEQ ID NO.1, its isoelectric point (pI) is 12.96, and its theoretical molecular weight (MW) is 3456.35Dalton.

[0030] The synthesized polypeptide is purified by reverse liquid phase high-pressure chromatography and identified by protein spectrum, and the purity is higher than 95%.

[0031] 2. Reversed-phase liquid chromatography column (C18) for deep purification of AIK polypeptide:

[0032] (1) Add 2 mL of acetonitri...

Embodiment 2AI

[0039] Embodiment 2 AIK inhibits tumor cell growth activity in vitro

[0040] 1. The cell lines used and their sources:

[0041] Cell lines 95C (low metastatic giant cell carcinoma of the lung), 95D (high metastatic giant cell carcinoma of the lung), HL-60 (acute promyelocytic leukemia), Hela (cervical cancer), HL-7702 (normal liver cells), B95 -8 (Epstein-Barr virus-infected leukemia), HO-8910PM (high metastatic ovarian cell carcinoma), HO-8910 (low metastatic ovarian cell carcinoma), SMMC-7721 (liver cancer cells), U2OS (myeloma cells), A549 (lung Adenocarcinoma cells) and other tumor cell lines were purchased from ATCC cell bank in the United States and stored in liquid nitrogen for a long time. The mouse liver cancer H22 ascites cell line is a cell bank of Harbin Medical University, and is now preserved by intraperitoneal inoculation of Kunming mice. All cells were cultured in DMEM medium (containing 10% FBS, purchased from Invitrogen) at 37°C, 5% CO 2 Incubator subcult...

Embodiment 3AI

[0056] Example 3 AIK inhibits the growth of subcutaneously transplanted tumors in tumor-bearing mice

[0057] 1. Experimental animals: 4-5 weeks old, 20-25g male Kunming mice, provided by the Experimental Animal Center of Harbin Medical University. The operation of animal experiments complied with the relevant rules of animal experiments of Harbin Medical University.

[0058] 2. Establishment of H22-bearing liver cancer cell subcutaneous xenograft mouse model

[0059] The mouse liver cancer H22 cells frozen in liquid nitrogen were quickly thawed in a 37°C water bath, and 3 Kunming mice were inoculated intraperitoneally, 0.5ml / mouse. One H22 ascites tumor model mouse was taken on the 8th day of passage, and sacrificed by cervical dislocation, and 8 ml of milky white ascites in the abdominal cavity of the mouse was sucked aseptically in an ultra-clean bench. 1:10 dilution and counting, PBS adjusted the concentration of tumor cell suspension to 5×10 6 pieces / ml. Disinfect the...

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Abstract

The invention discloses an artificially synthesized cationic peptide (named as AIK) and applications thereof in preparing an anti-tumor drug. The amino acid sequence of the cationic peptide is shown as SEQ ID NO.1. A research shows that the AIK has an obvious inhibition effect on various solid tumor cells, especially leukemic cells, cells of pulmonary giant cell cancer and hepatoma ascites cells, the anti-tumor activity is possibly combined an anionic acceptor on the surface of tumor cells so as to induce the tumor cells to be subjected to apoptosis and necrosis, and the anti-tumor activity is higher than that of the anti-tumor peptide molecules reported by other researchers. More importantly, the artificially synthesized 25-peptide drug molecules have higher specificity on the tumor cells. After the anti-tumor drug is locally administrated by subcutaneous tumor-bearing mice, the subcutaneous growth of liver cancer cells H22 of the tumor-bearing mice can be obviously inhibited and toxic and side effects are lower than chemotherapeutic drugs amethopterin. The artificially synthesized cationic peptide can provide theoretical basis and experimental materials for further researching and developing polypeptide anti-tumor new drugs and small-size targeted anti-tumor new drugs.

Description

technical field [0001] The invention relates to an artificially synthesized cationic peptide and its use in preparing new anticancer drugs, in particular to an artificially synthesized cationic 25 peptide and its use in inducing tumor cell apoptosis. The invention belongs to the field of biotechnology. Background technique [0002] Malignant tumors are a type of disease that seriously threatens human health and life. Although the existing anti-tumor drugs have certain curative effects on most tumors, they still have low treatment efficiency, poor selectivity, large toxic and side effects, and easy production of tumor cells. issues of drug resistance. Therefore, it is still an urgent task to find anti-tumor drugs with high efficiency, low toxicity and strong specificity from different ways. Anti-tumor polypeptide drugs have small molecular weight (generally less than 50 amino acids), high activity, easy penetration and absorption, and low immunogenicity. Its structure is s...

Claims

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Application Information

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IPC IPC(8): C07K14/00C12N15/11A61K38/16A61P35/00
CPCA61K38/00C07K14/00
Inventor 周春水范芳芳徐晖孙慧莹傅松滨白静刘佳玮张海员宁雪莲李亦兰
Owner HARBIN MEDICAL UNIVERSITY
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