PEGylation polypeptide with tumor inhibition function, and preparation method and application thereof

A peptide and tumor technology, applied in the biological field, can solve the problems of low solubility and high immunogenicity, and achieve the effects of inhibiting tumor growth, prolonging residence time, and inhibiting tumor cell proliferation.

Inactive Publication Date: 2018-11-23
BEIJING COMINGHEALTH BIO TEC CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Although peptide drugs have a specific action site and definite curative effect, their clinical application is severely restricted by defects such as low solubility, high immunogenicity, easy degradation by proteases, and clearance by the kidney.

Method used

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  • PEGylation polypeptide with tumor inhibition function, and preparation method and application thereof
  • PEGylation polypeptide with tumor inhibition function, and preparation method and application thereof
  • PEGylation polypeptide with tumor inhibition function, and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] Embodiment 1, PEGylated polypeptide mPEG 2000 Preparation of CY

[0064] (1) Obtaining peptides

[0065] The polypeptide obtained by adding cysteine ​​to the N-terminal of PIBC shown in SEQ ID NO.1 in the sequence listing is denoted as CY, and the amino acid sequence of CY is shown in SEQ ID NO.2 in the sequence listing, provided by Jill Biochemical (Shanghai) Co., Ltd. synthesized the polypeptide CY shown in SEQ ID NO.2.

[0066] (2) PEGylation of polypeptides

[0067] Raw materials: mPEGxMal (methoxy polyethylene glycol maleimide, x is the average molecular weight of PEG; Mal means maleimide, and maleimide is modified at one end of PEG; m means methoxy, form Oxygen is attached to the other end of PEG). Its chemical structural formula is shown in formula V:

[0068]

[0069] The molecular formula of PEG is HO(CH 2 CH 2 O) nH, the definition of n in formula I and formula V is the same as that of n in the PEG molecular formula. The mPEGxMal (x=2000, the averag...

Embodiment 2

[0076] Embodiment 2, PEGylated polypeptide mPEG 5000 Preparation of CY

[0077] The mPEGxMal (x=5000, the average molecular weight of PEG is 5000, mPEG 5000 Mal, a product of Beijing Jiankai Technology Co., Ltd.) undergoes an addition reaction with the sulfhydryl group of the N-terminal amino acid cysteine ​​of polypeptide CY through a highly chemoselective Michael addition reaction to obtain mPEG 5000 Cy.

[0078] mPEG 5000 CY is prepared according to the method of 1 in step (two) in Example 1, mPEG 2000 Mal is replaced by mPEG 5000 Mal, react until the peptide reaction is complete.

[0079] Agilent 1200 reversed-phase high-performance liquid chromatography was used to purify the reaction product obtained according to the above steps. Chromatographic column model: Angilent Eclipse XDB-C18 Semi-Prep, 5μm, 9.4×250mm. Chromatographic operating conditions: linear gradient elution, the eluent is composed of A2 solution and B2 solution in Example 1. Linear gradient elution:...

Embodiment 3

[0082] Embodiment 3, PEGylated polypeptide mPEG 2000 Preparation of LC

[0083]The polypeptide obtained by adding cysteine ​​to the C-terminus of PIBC shown in SEQ ID NO.1 in the sequence listing is denoted as LC, and the amino acid sequence of LC is shown in SEQ ID NO.3 in the sequence listing, provided by Jill Biochemical (Shanghai) Co., Ltd. synthesized the polypeptide LC shown in SEQ ID NO.3.

[0084] The mPEGxMal (x=2000, the average molecular weight of PEG is 2000, mPEG 2000 Mal, a product of Beijing Jiankai Technology Co., Ltd.) through a highly chemoselective Michael addition reaction and the sulfhydryl group of the C-terminal amino acid cysteine ​​of the polypeptide LC undergo an addition reaction to obtain mPEG 2000 LC, mPEG 2000 The specific process of preparation, purification and characterization of LC is the same as that described in step (2) in Example 1, except that the polypeptide CY in Example 1 is replaced with polypeptide LC to obtain the fluffy PEGylate...

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Abstract

The invention discloses a PEGylation polypeptide with a tumor inhibition function, and a preparation method and application thereof. The PEGylation polypeptide is obtained after modification of a polypeptide named PIBC by PEG. A compound A and a compound B are subjected to Michael addition reaction to form a carbon sulphur single bond to achieve modification. The compound A is PEG with one end modified by maleimide, and the compound B is PIBC, wherein a sulfydryl cysteine residue is introduced to the end N or C of PIBC. PIBC is the polypeptide shown as SEQ ID No. 1. The end-N amino acid residue of PIBC takes the first place of the SEQ ID No. 1. The Michael addition reaction occurs between maleimide and sulfydryl. The PEGylation polypeptide has the advantages that the inhibition effect of PIBC on the activity of heat shock protein gp96 is maintained, half-life in vivo of PIBC is prolonged, and the PEGylation polypeptide has potential value for development of new drugs.

Description

technical field [0001] The invention relates to a PEGylated polypeptide with the function of inhibiting tumors and its preparation method and application in the field of biotechnology. Background technique [0002] According to the "2015 China Cancer Statistics Report" published by the National Cancer Registry Center, breast cancer is the type of cancer with the highest incidence rate among Chinese women, and its incidence rate is increasing at a rate of 3% per year, making it the fastest-growing mortality rate in cities. cancer. Currently, the main treatment methods for breast cancer include surgery, chemotherapy, endocrine therapy and targeted therapy. Targeted therapy has the characteristics of accurate positioning, strong pertinence, and low toxicity. It can significantly prolong the survival period of patients and improve the quality of life of patients. It is especially suitable for advanced patients or patients who cannot tolerate radiotherapy and chemotherapy. The ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/00A61K38/16A61P35/00
CPCA61K38/00C07K14/00C07K19/00A61K38/16A61P35/00
Inventor 胡俊孟颂东杨哲杨博吴飚丁筠
Owner BEIJING COMINGHEALTH BIO TEC CO LTD
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