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18F click labelling transferrin receptor targeting polypeptide T7 as well as preparation method and application thereof

A transferrin, targeting peptide technology, applied in the preparation methods of peptides, chemical instruments and methods, peptides, etc., can solve the problems of long time, unfavorable automatic synthesis, low radioactive recovery rate, etc., and achieve radiochemical yield. High, the effect of reducing one purification process and reducing the time of synthesis and purification

Active Publication Date: 2016-11-09
GUANGZHOU GENERAL HOSPITAL OF GUANGZHOU MILITARY COMMAND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

According to the method reported in the literature, the preparation of polypeptide radioactive probes can only be completed after two HPLC purifications. The radioactive recovery rate is not high, and it takes a long time, which is not conducive to automatic synthesis.

Method used

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  • 18F click labelling transferrin receptor targeting polypeptide T7 as well as preparation method and application thereof
  • 18F click labelling transferrin receptor targeting polypeptide T7 as well as preparation method and application thereof
  • 18F click labelling transferrin receptor targeting polypeptide T7 as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 118

[0049] Example 1 18 Chemical Synthesis of F-TEG-T7

[0050] 1) Synthesis of labeled precursor TsOTEGay

[0051] At 0°C, slowly add 60% NaH (2g, 0.050mol) into a THF (60mL) solution of TEG (11.6g, 0.077mol) and stir. After no bubbles emerge, add 3-bromopropyne ( 2.1 mL, 0.039 mol), the reaction was followed by TLC (petroleum ether: ethyl acetate = 1:2). The solution changed from colorless to brown, and the reaction was completed. The reaction solution was centrifuged, the solvent was concentrated, and purified by column chromatography with petroleum ether: ethyl acetate = 1:2-1:4. The yellow liquid product (TEGay) was obtained in 3.58 g, the yield was 49%.

[0052] Dissolve TEGay (2.00 g, 0.010 mol) p-hydroxybenzoyl chloride (0.012 mol) in 50 mL of anhydrous CH 2 Cl 2 , cooled to 0°C. Add 2.98g (0.053mol) KOH solid in 10 times, and stir at 0°C for 1h. Stir at room temperature for another 5 h (TLC to track the reaction, petroleum ether: ethyl acetate = 2:1). After the re...

Embodiment 2

[0058] Example 2 Standards 19 Chemical Synthesis of F-TEG-T7

[0059] H-TEGay (100 mg, 0.5 mmol) was dissolved in 2.5 mL of anhydrous dichloromethane and cooled to 0 °C. DAST (132 μL, 1 mmol) was added slowly, and after reacting for 1 h at 0° C., the reaction was continued at room temperature for 5 h. After the reaction, the product was separated with a silica gel column (eluent: ethyl acetate:petroleum ether=1:6~1:3), and the light yellow oily product was obtained after drying the solvent.

[0060] The collected product was characterized by NMR spectroscopy as 19 F-TEGay, for second-step click reaction synthesis of standards 19 F-TEG-T7. 19 F-TEGay (0.34mg, 1.79μmol) was dissolved in 400μL acetonitrile, T7 polypeptide (2mg, 1.79μmol), CuSO 4 .5H 2 O (2.2mg, 8.9μmol) and ASC (14.2mg, 71μmol) were dissolved in 800μL H 2 In O, mix acetonitrile and aqueous solution, seal, and react at 60°C for 30min. After the reaction, HPLC was used for detection and separation, and afte...

Embodiment 3

[0067] Example 3 18 Lipid-water partition coefficient experiment of F-TEG-T7

[0068] Take 100μL 18 The F-TEG-T7PBS preparation was placed in a 1.5mL centrifuge tube (containing 500μL n-butanol and 400μL PBS), sealed, vortexed at room temperature for 2min, and then centrifuged at high speed for 3min (10000r / min) until the two phases were balanced. Use a pipette gun to sample 100 μL each from the organic phase and the aqueous phase and place them in two gamma counter tubes to determine the gamma count. Repeat the sampling measurement 3 times. The average lg D is calculated according to the following formula 7.4 value.

[0069] d 7.4 =lg(N 1 / N 2 ); where: N 1 , radioactive count rate per ml n-octanol, min -1 ·mL -1 ; N 2 , radioactive count rate per ml buffer solution PBS, min -1 ·mL -1 .

[0070] Calculated by measurement, 18 The lipid-water partition coefficient of F-TEG-T7 is -0.51±0.01, and the targeting polypeptide tends to be more hydrophilic. It can be pre...

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Abstract

The invention discloses 18F click labelling transferrin receptor targeting polypeptide T7 as well as a preparation method and application thereof. According to the preparation method, synthesis steps of click chemistry are improved, addition amount of a prosthetic group TsOTEGay is reduced, the addition amount of polypeptide T7 is appropriately increased, feeding ratio of the prosthetic group to the polypeptide T7 is adjusted to be 1:(1-2); after 18F-TEGay is synthesized through reaction, a polypeptide precursor is directly added for carrying out click reaction without carrying out HPLC purification, a 'one-pot two-step' process is adopted for synthesizing 18F-TEG-T7, and finally HPLC purification is carried out; and one purification process is reduced, the whole synthesis and purification time is greatly reduced and can be controlled to be within 120 minutes, radiochemical yield is about 28.26 minus and plus 7.43% (n=3). The optimized 'two-step one-pot' click reaction can be applied to research of 18F labelling polypeptide, and a novel tumor targeted probe can be prepared.

Description

technical field [0001] The present invention specifically relates to 18 F click marker transferrin receptor targeting polypeptide T7 and its preparation method and application. Background technique [0002] Glioma is one of the most malignant brain tumors, originating from glial cells, and is the most common primary central nervous system tumor, accounting for about 42% of all brain tumors. The highly proliferative, invasive and aggressive nature of glioma makes its treatment still quite difficult. Timely and early accurate diagnosis of glioma is the key to treatment. [0003] Transferrin Receptor (TfR) is highly expressed on the surface of tumor cells, especially glioma cells and brain capillary endothelial cells. T7 polypeptide (sequence HAIYPRH, His-Ala-Ile-Tyr-Pro-Arg-His) is a targeting polypeptide screened from cells expressing human transferrin receptor TfR by using phage display technology. The role of TfR. In the in vitro cell experiment, the T7 polypeptide has...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/06C07K1/13A61K51/08A61K49/04A61K47/42A61P35/00
CPCA61K47/42A61K49/04A61K51/08C07K7/06C07K19/00
Inventor 王欣璐尹吉林黄伟耀
Owner GUANGZHOU GENERAL HOSPITAL OF GUANGZHOU MILITARY COMMAND
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