Human interleukin 2-polyethylene glycol conjugate and application thereof

A technology of interleukin and polyethylene glycol, which is applied in the field of human interleukin 2-polyethylene glycol conjugates, can solve the problems of weakened binding ability, complicated mechanism of action, and no performance, so as to reduce the binding activity , high product uniformity, and the effect of reducing the frequency of administration

Active Publication Date: 2021-11-26
NOVOCODEX BIOPHARMACEUTICALS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] 1. Simple amino acid site-directed mutation can reduce the binding ability to IL-2Rα, or enhance the binding ability to IL-2Rβ or IL-2Rγ, but it cannot effectively prolong the half-life of the molecule, and the mutant product is prone to immunogenic reactions in vivo , it is easy to reduce the biological activity of the product and cause a greater risk of toxicity
[0008] 2. Although simple fusion expression (such as fusion with Fc or HSA) or IL-2 modification can prolong the half-life of the molecule, it has no obvious advantage in actual use compared with unmodified IL-2
Fusion expression can only realize the modification of the fusion molecule at the N-terminus or C-terminus of the target protein, and cannot optimize the modification site
[0009] 3. Partial site-directed mutation combined with fusion expression of IL-2 does not show special advantages in practical application
[0010] 4. Conventional non-site-specific coupling of PEGylated IL-2 does not show special advantages in practical applications. PEGylated IL-2 that comprehensively weakens the binding ability of IL-2Rα has achieved staged success, but the non-site-specific coupling process The characteristics make it difficult to control the production process and quality, the molecular structure is complicated, and the mechanism of action is complicated.
However, this coupling method and chemical modification method need to introduce a relatively expensive alkyne structure, and an acceptable drug-antibody coupling ratio can only be obtained when the equivalent is used, which increases the corresponding production cost and process. The process is also more complex and the process conditions are harsh

Method used

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  • Human interleukin 2-polyethylene glycol conjugate and application thereof
  • Human interleukin 2-polyethylene glycol conjugate and application thereof
  • Human interleukin 2-polyethylene glycol conjugate and application thereof

Examples

Experimental program
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preparation example Construction

[0079] The reagents or raw materials used in the preparation examples and examples of the present invention are all commercially available products unless otherwise specified; the experimental methods used are all conventional methods in the art unless otherwise specified.

[0080] Human YT cells in the literature "Yodoi, J. et al. (1985). TCGF(IL 2)-receptor inducing factor(s). I. Regulation of IL 2 receptor on a natural killer-like cell line (YTcells). Journal of Immunology, 134(3), 1623-1630", the public can obtain from Zhejiang Xinma Biomedical Co., Ltd.

preparation example 1

[0081] Preparation Example 1 The preparation of unnatural amino acid NBOK

[0082] The structural formula of NBOK is as follows:

[0083]

[0084] The reaction process is shown in the figure below:

[0085]

[0086] The preparation process includes the following steps:

[0087] a) In the reaction flask, add p-methylacetophenone (4.0mL, 30.0mmol), add solvent DCE (50.0mL), add NBS (6.41g, 36.0mmol) and BPO (0.05g, 0.3mmol), the mixture After reflux at 80°C for 24 hours, the container was cooled in ice water, and a solid precipitated, which was removed by filtration and washed with saturated Na 2 CO 3 Washed 3 times, extracted 3 times with DCM, combined the organic phases, added anhydrous sodium sulfate to dry, filtered, and concentrated under reduced pressure to obtain the crude product 1-1 (5.46 g, yield 85%), which was directly used in the next step without purification .

[0088] b) In a reaction flask, add product 1-1 (2.73g, 12.80mmol), add solvent dioxane (40mL...

preparation example 2

[0093] Preparation Example 2 The preparation of unnatural amino acid NPAK

[0094] The structural formula of NPAK is as follows:

[0095]

[0096] The reaction process is shown in the figure below:

[0097]

[0098] The preparation process includes the following steps:

[0099] a) In the reaction flask, add p-chloroacetophenone (1.00g, 6.47mmol), under nitrogen atmosphere, add diethyl malonate (6.84g, 47.70mmol), KHCO 3 (0.97g, 9.70mmol) and K 2 CO 3 (1.34g, 9.70mmol), then add Pd(dba) 2 (0.019g, 0.030mmol) and P(t-Bu) 3 HBF 4 (0.021g, 0.071mmol), after the addition was completed, the nitrogen was replaced, and the temperature was raised to 160°C for 40h. After the reaction was complete as detected by TLC, water (30 mL) was added to the reaction solution, extracted 3 times with EA, the organic phases were combined, washed 2 times with water, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure at 0-5°C to obtain a colorless The t...

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Abstract

The invention provides a human interleukin 2-polyethylene glycol conjugate and application thereof. The human interleukin 2-polyethylene glycol conjugate provided by the invention comprises recombinant human interleukin 2 containing at least one non-natural amino acid and PEG coupled to the at least one non-natural amino acid, the non-natural amino acid is a carbonyl terminal group-containing compound with a structure as shown in a formula (I) or an enantiomer thereof, and PEG is coupled to the at least one non-natural amino acid by forming an oxime bond between the carbonyl end group and PEG containing a hydroxylamine end group. The human interleukin 2-polyethylene glycol conjugate provided by the invention can be singly used or combined with other anti-tumor drugs, and is used for treating diseases such as solid tumors and hematologic tumors.

Description

technical field [0001] The invention relates to the field of biopharmaceuticals, in particular to a human interleukin 2-polyethylene glycol conjugate and an application thereof. Background technique [0002] Interleukin-2 (Interleukin 2, IL-2, often referred to as Interleukin 2) is an important immune regulatory factor produced by activated type I helper T lymphocytes (Th1), once known as T cell growth factor, Its main biological function is to promote T cells (including CD4 + and CD8 + T cells) growth, proliferation, differentiation, and promote the further secretion of cytokines. In addition, interleukin 2 also stimulates the proliferation of NK cells, enhances NK killing activity and produces cytokines, induces the production of LAK cells; promotes the proliferation of B cells and secretes antibodies; Important role (Gaffena S.L., Cytokine 28:109e123, 2004). [0003] Since the first discovery of IL-2 by Morgan et al. in 1976, it has been widely used clinically. In 1...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/55A61K38/20A61K47/60A61P35/00A61P35/02A61P37/04
CPCC07K14/55A61K38/2013A61K47/60A61P35/00A61P35/02A61P37/04Y02A50/30
Inventor 梁学军叶诚浩夏钢陈龙飞霍鹏超杨金纬应跃斌宫丽颖林欣巩尊洋衡新朱小愚朱丽飞黄浩叶谋田丁文焦琳祝静静
Owner NOVOCODEX BIOPHARMACEUTICALS CO LTD
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