Preparation method for poly(ethylene glycol) modified recombinant human interleukin-2

A technology of polyethylene glycol and human interleukin, applied in the field of preparation of recombinant human interleukin 2, can solve the problems of poor uniformity of coupling products and large loss of activity, etc., achieve high antiviral activity, simple steps, easy quality control and scale-up The effect of production

Inactive Publication Date: 2013-07-10
SHENZHEN YATAIXING IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] The purpose of the embodiment of the present invention is to provide a preparation method of polyethylene glycol-modified recombinant human interleukin 2,...

Method used

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Examples

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preparation example Construction

[0015] The embodiment of the present invention provides a preparation method of polyethylene glycol-modified recombinant human interleukin-2, comprising the following steps:

[0016] S01 activates mPEG with an activator to obtain mPEG activated molecules;

[0017] S02 reacts the mPEG activated molecule with recombinant human interleukin 2 in a buffer solution with a pH of 7.5-9.0, then adds a glycine solution to terminate the reaction, and separates and purifies the reaction product.

[0018] Specifically, the monomethoxypolyethylene glycol (Methoxy Polyethylene Glycol), referred to as mPEG, used in step S01 is a kind of polyethylene glycol, which has good water solubility, wettability, lubricity, physiological inertness, Non-irritating to the human body, mild in nature.

[0019] In a preferred embodiment, the molecular weight of mPEG used in step S01 is between 10kDa~100kDa, more preferably between 10kDa~50kDa, more preferably 10kDa~30kDa.

[0020] In this step S01, the act...

Embodiment 1

[0038] The molecular weight of mPEG used in this example is 10 kDa, and the activator is succinic anhydride and N-hydroxysuccinimide. The specific operation steps are as follows:

[0039] 1. mPEG activation

[0040] Dissolve mPEG in dioxane, add an appropriate amount of succinic anhydride so that the molar amount of succinic anhydride is slightly more than that of mPEG, use pyridine as a catalyst, and reflux at 100°C for 4 hours. After the reaction solution was cooled, anhydrous diethyl ether was added under vigorous stirring until the precipitation no longer appeared, and then stirred in an ice bath for 30 min. After filtration, the resulting precipitate was dissolved in dichloromethane, and the insoluble matter was removed by filtration again. Anhydrous ether was added to the filtrate until no precipitation occurred, and then stirred in an ice bath for 15 min. The precipitate was filtered again and dried in vacuo. The dry product and N-hydroxysuccinimide were dissolved in...

Embodiment 2

[0046] The molecular weight of mPEG used in this example is 20kDa, and the activator is succinic anhydride and N-hydroxysuccinimide. The specific operation steps are as follows:

[0047] 1. mPEG activation

[0048] Dissolve mPEG in dioxane, add an appropriate amount of succinic anhydride to make the molar amount more than that of mPEG, use pyridine as a catalyst, and reflux at 0°C for 24h. After the reaction solution was cooled, anhydrous diethyl ether was added under vigorous stirring until the precipitation no longer appeared, and then stirred in an ice bath for 30 min. After filtration, the resulting precipitate was dissolved in dichloromethane, and the insoluble matter was removed by filtration again. Anhydrous ether was added to the filtrate until no precipitation occurred, and then stirred in an ice bath for 15 min. The precipitate was filtered again and dried in vacuo. The dry product is mixed with N-hydroxysuccinimide and dissolved in dimethylformamide (DMF), and th...

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Abstract

The invention provides a preparation method for poly(ethylene glycol) modified recombinant human interleukin-2, belonging to the field of biological medicine. The method comprises the following steps: activating mPEG with an activator so as to obtain an activated mPEG molecule; and reacting the activated mPEG molecule with recombinant human interleukin-2, adding a glycine solution to terminate a reaction and separating and purifying an obtained product; wherein the activator is one selected from the group consisting of succinic anhydride and N-hydroxysuccinimide, p-toluenesulfonic acid-chlorine, N,N'-carbonyl diimidazole, N,N'-disuccinimidocarbonate, p-nitrophenyl carbonate, benzotriazole carbonate, phenylsuccinimide carbonate and N-acetoxysuccinimide. The preparation method provided by the invention is simple and convenient to operate and is easy for quality control and enlarged production.

Description

technical field [0001] The invention belongs to the field of biomedicine, in particular to a preparation method of polyethylene glycol-modified recombinant human interleukin-2. Background technique [0002] Interleukin (interleukin), the abbreviation of interleukin, refers to the lymphokine that interacts between white blood cells or immune cells. It belongs to cytokines like blood cell growth factor. The two coordinate and interact with each other to complete hematopoietic and immune regulation functions. Interleukins play an important role in transmitting information, activating and regulating immune cells, mediating T and B cell activation, proliferation and differentiation, and inflammatory responses. [0003] Interleukin-2 (interleukin-2, IL-2), is a member of the interleukin family, also known as T cell growth factor, mainly produced by activated CD4+ T cells and CD8+ T cells, its It has a wide range of biological activities and can act on all T cell subsets. Recomb...

Claims

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Application Information

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IPC IPC(8): C07K14/55C07K1/107
Inventor 徐东叶垚平
Owner SHENZHEN YATAIXING IND
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