Coupling reagent with azaphenanthrone structure and application of coupling reagent in preparation of polypeptide and protein conjugate

A protein, conjugation technology

Active Publication Date: 2021-11-23
NANJING UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the modification method of lysine residues in polypeptides or proteins is relatively simple. The classic modification method is to use pre-prepared N-hydroxysuccinimide (NHS) esters to form stable amide bonds with lysine residues.
However, NHS esters are easily hydrolyzed in aqueous solution and will react with tyrosine, serine and threonine to form by-products, which limits its application.

Method used

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  • Coupling reagent with azaphenanthrone structure and application of coupling reagent in preparation of polypeptide and protein conjugate
  • Coupling reagent with azaphenanthrone structure and application of coupling reagent in preparation of polypeptide and protein conjugate
  • Coupling reagent with azaphenanthrone structure and application of coupling reagent in preparation of polypeptide and protein conjugate

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Experimental program
Comparison scheme
Effect test

Embodiment 1-8

[0040] The present invention synthesizes compound through embodiment 1-8:

[0041]

[0042] The synthetic route is:

(1)

[0043]

(2)

[0044]

(3)

[0045]

(4)

[0046]

[0047] Raw material 2a in embodiment 1,4 is synthesized by the following method:

[0048] synthetic route:

[0049]

[0050] The specific synthesis method is:

[0051] Add 5g sodium hydrogen (122mmol, 1.2eq.), 0.95g tetrabutylammonium iodide (5.1mmol, 0.025eq.) into a 500ml round bottom flask, N 2 For protection, add 300ml of anhydrous tetrahydrofuran, drop to 0°C, add dropwise 12ml of diethylene glycol monomethyl ether S2 (102mmol, 1.0eq.); stir at room temperature for 15min; drop to 0°C, add dropwise 10.5ml of propyne bromide S1 (122mmol, 1.2eq.), reaction at room temperature, TLC tracking, after the reaction, add 30ml of ice water to quench, concentrate under reduced pressure, spin off most of THF, extract with EA, combine organic phases, anhydrous Na 2 SO 4 Drying, concentrat...

Embodiment 2

[0052] Raw material 2b is synthesized by the following method among the embodiment 2:

[0053] synthetic route:

[0054]

[0055] The specific synthesis method is:

[0056] Add 12.6g sodium hydrogen (315mmol, 3.0eq.), 1.9g tetrabutylammonium iodide (5.25mmol, 0.05eq.) into a 1000ml round bottom flask, N 2 For protection, add 400ml of anhydrous tetrahydrofuran, drop to 0°C, add 10ml of diethylene glycol S3 (105mmol, 1.0eq.) dropwise; stir at room temperature for 30min; 3.0eq.), the drop was completed and moved to room temperature, followed by TLC. After the reaction was completed, 40ml of ice water was added to quench, concentrated under reduced pressure, and most of the tetrahydrofuran was spun off, extracted with EA, combined organic phases, anhydrous Na 2 SO 4 Drying, concentration under reduced pressure, and column chromatography (PE:EA=50:1-7:1) yielded 17.5 g of compound 2b with a yield of 91%.

[0057] Raw material 2c is synthesized by the following method in embo...

Embodiment 6

[0062] Raw material 2d is synthesized by the following method in embodiment 6:

[0063]

[0064] The specific synthesis method is:

[0065] 55.4mg compound S4 (0.67mmol, 1.0eq.), 216mg compound S6 (0.81mmol, 1.2eq.) was added in a 10ml round bottom flask, N 2 Protection, adding 0.3ml of anhydrous triethylamine, 6ml of anhydrous N,N-dimethylformamide, stirring at room temperature, TLC tracking, after the reaction, 10% hydrochloric acid adjusted to neutral, EA extraction, combined organic phase, no Water Na 2 SO 4 It was dried, concentrated under reduced pressure, and separated by column chromatography (PE:EA=200:1-15:1) to obtain 133 mg of compound 2d with a yield of 85%.

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Abstract

The invention belongs to the technical field of organic synthesis and protein modification, and particularly relates to a coupling reagent with an azaphenanthrone structure and application of the coupling reagent in preparation of polypeptide and protein conjugage. The polypeptide and protein coupling reagent with the azaphenanthrone structure is good in stability in an aqueous solution, and protein modification operation is simple and convenient. The coupling reagent is novel in structure and different from existing known modification reagents in structure, the coupling product is good in connection stability, and a new technology and a new mode are provided for various functional modifications of polypeptides and proteins.

Description

technical field [0001] The invention belongs to the technical field of organic synthesis and protein modification, in particular to a coupling reagent with an azaphilone structure and its application in preparing polypeptide and protein conjugates. Background technique [0002] The modification of polypeptide or protein is an important means to effectively improve its physical and chemical properties, such as increasing water solubility, prolonging the action time in vivo, and reducing toxic and side effects. The introduction of specific functional groups (directing groups, fluorophores, and drugs) can change the distribution of polypeptides or proteins in living systems, perform biomedical imaging, and treat diseases. Generally, coupling reagents need to react with some functional groups in polypeptides or proteins, and lysine is important due to its high content in proteins (5.9% in the human proteome) and is usually exposed on the surface of proteins modification site. ...

Claims

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Application Information

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Patent Type & AuthorityApplications(China)
IPC IPC(8): C07D311/76C07D405/12C07D405/14A61K47/54A61K38/08A61K38/47A61K38/46A61K38/10
CPCC07D311/76C07D405/12C07D405/14A61K47/545A61K38/08A61K38/47C12Y302/01017A61K38/465A61K38/10Y02P20/55
Inventor姚祝军伊善东王欢魏思媛奚婕
OwnerNANJING UNIV