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Tumor targeting polypeptide and preparation method thereof

A targeted, breast tumor technology, applied in the medical field to achieve the effects of improved targeting, easy synthesis and convenient use

Inactive Publication Date: 2012-02-29
昆明医学院第一附属医院
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  • Summary
  • Abstract
  • Description
  • Claims
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Problems solved by technology

There is no receptor-mediated carrier used in the study of affinity and specificity of tumors, and the use of receptor-mediated carriers to detect the affinity and specificity of breast cancer cells has not been reported at home and abroad

Method used

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  • Tumor targeting polypeptide and preparation method thereof

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Experimental program
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Effect test

Embodiment

[0029] Get MDA-MB-231 cells in the logarithmic growth phase and inoculate them on a 150mm tissue culture dish at 37°C for 12 hours, replace with fresh culture medium (L15 culture medium containing 10% FBS), and add Chloroquine to the culture medium to make the final concentration reach 100μM, 37℃, 30min; 3×10 11 TU’s pC89 phage peptide library phage was added to the culture dish, 8h-12h, 5min on ice; wash the cells 4 times with 40ml HBSS at 4°C, wash once with HBSS(-), and then treat the cells with 11ml Subtilisin for 1h to prepare the cell suspension , centrifuge to remove Subtilisin; then treat the cells with 6ml of HBSS(-) solution containing 1mM PMSF and 2mM EDTA for 15min, wash; of bacteriophages.

[0030] Transform the collected cell lysates into competent E. coli XLI-Blue, amplify on LB plates, collect the transformed bacteria liquid, add M13KO7 helper phages to prepare phage particles; add the amplified phages to the cells again for The next round of screening-amplif...

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Abstract

The invention relates to a tumor targeting polypeptide and a preparation method thereof. The amino acid sequential structure of the polypeptide of the invention is CFPVPGHDLVC. The preparation method comprises the following steps: co-culturing a pC89 phage peptide library for expressing 108 protein polypeptides with different sequences and human breast cancer cell strain MDA-MB-231 repeatedly formany times; screening phages which express specific polypeptides and can penetrate a cell membrane and enter cytoplasm and / or nucleus, amplifying the phages in vitro and carrying out DNA sequencing, and deducing exogenous amino acid sequences inserted on the phages; co-culturing the screened specific polypeptide phages, other human tumor cells and normal cells in vitro, and detecting the specificity of cell tumor of the polypeptide phages; and according to the polypeptide sequences and the detecting result of the cell specificity, artificially synthesizing the tumor targeting polypeptide. Theinvention has potential clinical application values for specific vectors of breast cancer targeting gene therapy, and also provides powerful technical support for screening polypeptides with affinityand specificity of other types of malignant tumor cell strains in the future. In addition, the polypeptide of the invention contains only 9-11 amino acid residues, can be synthesized easily, and has relatively small spatial position change, easy quality control of the polypeptide and convenient use.

Description

technical field [0001] The invention relates to the field of medical technology, in particular to a tumor-targeting polypeptide and a preparation method thereof. Background technique [0002] Overcoming tumors has always been the focus of biomedical circles. Over the past hundred years, three major clinical treatment methods for tumors have been developed: surgery, radiotherapy, and chemotherapy. These treatments prolong the survival period of patients with tumors to a certain extent, but in addition to resecting and destroying tumors, they also damage the structure and function of normal tissues during clinical application, especially when chemotherapy drugs are used systemically, the drugs pass through the blood The circulation reaches various tissues of the human body and also exerts a killing effect on normal cells, causing systemic adverse reactions and seriously affecting the final effect of treatment. Therefore, the research and development of new tumor treatment me...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/06G01N33/68A61K49/00
Inventor 董坚陈明清刘流刘为青张克健
Owner 昆明医学院第一附属医院
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