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Aminopeptidase N inhibitor and preparation method and application thereof

An inhibitor, aminopeptidase technology, applied in the chemical field, can solve the problems of poor metabolic stability, weakening the ability of macrophages and NK cells to recognize and kill tumor cells, and decreased immunity of the body to achieve strong anti-tumor activity Effect

Inactive Publication Date: 2014-06-11
WEIFANG HIGH TECH BIOLOGICAL PARK DEV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

2) APN participates in T lymphocyte-dependent inflammatory responses, can be expressed on the surface of antigen-presenting cells, and degrades immune active substances (such as interleukin-8); it also reduces the ability of T cells to recognize their antigens, and at the same time weakens macrophages Cells and NK cells have the ability to recognize and kill tumor cells, reducing the body's immunity
[0005] The patent (CN200910020656.9) reported α-aminoacylcycloimide peptide metalloprotease inhibitors and their applications. Among them, compound 13f has strong anti-tumor activity in vivo and in vitro, but the structure of compound 13f contains Degraded peptide bonds, poor metabolic stability in vivo

Method used

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  • Aminopeptidase N inhibitor and preparation method and application thereof
  • Aminopeptidase N inhibitor and preparation method and application thereof
  • Aminopeptidase N inhibitor and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1. Synthesis of compound of the present invention (I)

[0025] 1) Preparation of (S)-tert-butyl-2,6-dioxopiperidin-3-ylcarbonamide (2)

[0026] Boc-L-glutamine (9.86g, 40.0mmol) and N-hydroxysuccinimide (4.6g, 40.0mmol) were dissolved in 100ml of tetrahydrofuran (THF), and 50ml containing di Cyclohexylcarbodiimide (8.24g, 40mmol) in THF. After about 1 hour of dripping, the ice bath was removed, and after stirring at room temperature for 3 hours, the reaction was refluxed for 10 hours. The reaction solution was cooled to room temperature, and the solvent was distilled off under reduced pressure. The residue was added to 40 ml of ethyl acetate and concentrated again. After the final residue was added with 50 ml of ethyl acetate, it was kept in the refrigerator overnight. Filter with 2.0g of diatomaceous earth, wash the filtrate with water (20ml×1) and saturated brine (20ml×1), dry over anhydrous magnesium sulfate, filter, and concentrate to dryness. Then r...

Embodiment 2

[0045] Example 2. Activity experiment of target compound inhibiting aminopeptidase N (In vitro)

[0046] The assay for aminopeptidase N (APN) inhibitory activity is described in Lejczak, B et al. Biochemistry, 1989, 28, 3549. The substrate L-leucyl-p-nitroaniline is degraded by APN to produce p-nitroaniline which absorbs at 405nm, and the concentration of p-nitroaniline is positively correlated with the enzyme activity. The content of p-nitroaniline is determined by detecting the absorbance at 405nm, so as to determine the activity of aminopeptidase, which indirectly reflects the degree of inhibition of the enzyme activity by the inhibitor.

[0047] For the specific method and operation steps, refer to CN100560568C "Cyclic imide peptide metalloprotease inhibitor and its application". The experimental results are shown in Table 1.

[0048] Table 1 In vitro enzyme inhibitory activity test results of target compounds

[0049]

[0050] The value in the table is the average v...

Embodiment 3

[0052] Example 3. The target compound inhibits tumor cell proliferation activity test (In vitro)

[0053]The in vitro anti-proliferation test of the compound on tumor cells was carried out using the thiazolium detection method (MTT method), and the cell suspensions of human ovarian cancer cell line (ES-2) or human breast cancer cell line (MDA-MB-231) were inoculated in In a 96-well plate, add medium containing different concentrations of compounds to each well, after incubation, stain with MTT, after continued incubation, measure the absorbance (OD value) of each well at 570nm on a microplate reader, and calculate the cell growth Inhibition rate to determine the activity of the compound.

[0054] 1. [Materials] ES-2 cell line, MDA-MB-231 cell line, tetramethylazoblue MTT, 10% fetal bovine serum, 96-well plate.

[0055] 2. [Method]

[0056] Cell culture The ovarian clear cell carcinoma ES-2 cell line and the breast cancer MDA-MB-231 cell line were cultured conventionally. Ce...

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Abstract

The invention discloses an aminopeptidase N inhibitor which has a chemical name of 2-((S)-3-(3-((S)-2-amino-3-phenylpropyl)urea)-2,6-dioxopiperidine-1-yl)-N-hydroxyacetamide hydrochloride and is a compound with a structural formula (I) described in the specification. The invention further discloses a preparation of the compound and application of the compound in preparation of antitumor medicines. The aminopeptidase N inhibitor obtained by the invention is matched with an active site of aminopeptidase N in space, thereby showing stronger antitumor activity in evaluation of in vitro and vivo antitumor activity evaluation.

Description

technical field [0001] The invention relates to a preparation method of an aminopeptidase N inhibitor and its medicinal use in antitumor, and belongs to the field of chemical technology. Background technique [0002] Aminopeptidase N (APN, CD13) usually refers to human alanyl aminopeptidase (EC3.4.11.2), which is a zinc ion-dependent hydrolytic protease belonging to the M1 family of metalloproteinases and homodimer The form exists on the cell membrane, which is the same substance as the cell surface antigen CD13; aminopeptidase N is expressed on the surface of various cells in various tissues, especially in the enterocytes of the small intestine and the epithelial cells of the proximal convoluted tubule of the kidney It is most abundant in , and is also expressed in the liver, placenta and uterus. Aminopeptidase N can hydrolyze and release N-terminal neutral amino acids or basic amino acids (Ala>Phe>Tyr>Leu>Arg>Thr>Trp) from peptides, amides or aromatic am...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D211/88A61K31/45A61P35/00
CPCY02P20/55C07D211/88
Inventor 张晓攀马文泉徐文方刘刚
Owner WEIFANG HIGH TECH BIOLOGICAL PARK DEV
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