Prolyl hydroxylase small molecular fluorescent probes and preparation method thereof

A technology of prolyl hydroxylase and fluorescent probe, which is applied in chemical instruments and methods, fluorescence/phosphorescence, analytical materials, etc., can solve the problems of complex test system, loss of binding ability of PHD enzyme, and decrease of binding ability of HIF fluorescent peptide and other problems, to achieve the effect of simple test system and optimized structure

Inactive Publication Date: 2019-02-01
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are still some defects in this method. For example, the system relies on 2-OG, because without 2-OG, the binding force of the HIF fluorescent peptide to the PHD enzyme is greatly reduced; in addition, the endogenous cofactor Fe in the system 2+ was replaced by Mn 2+ To avoid the HIF fluorescent peptide being hydroxylated and losing its binding ability to PHD enzymes
Therefore, the test system is still relatively complicated, and the Mn 2+ does not really reflect the relationship between the compound and the endogenous Fe-containing 2 + Binding of PHD enzymes

Method used

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  • Prolyl hydroxylase small molecular fluorescent probes and preparation method thereof
  • Prolyl hydroxylase small molecular fluorescent probes and preparation method thereof
  • Prolyl hydroxylase small molecular fluorescent probes and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Preparation of fluorescent probe Ⅰ-1

[0065] (3-((2-(2-azidoethoxy)-5-fluorophenyl)amino)propyl)carbamate tert-butyl ester (0.706g, 2.0mmol), (5-ethynyl-3- Hydroxypicolilyl) glycine (0.440g, 2.0mmol), tetrabutylammonium fluoride (1.0mol / L solution in THF, 1.56g, 6.0mmol), cuprous iodide (0.023g, 0.12mmol) and N, N-Diisopropylethylamine (1.0mL) was dissolved in MeOH (8mL) under N 2 The mixture was stirred at 80° C. for 5.5 hours under protection, and the reaction mixture was filtered and concentrated. The product was purified by column chromatography on silica gel (eluent: 30-50% EtOAc in petroleum ether) to afford (5-(1-(2-(2-((3-((tert-butoxycarbonyl)amino) ) Propyl) amino)-4-fluorophenoxy) ethyl)-1H-1,2,3-triazol-4-yl)-3-hydroxypyridinecarbonyl)glycine 0.620g; Soluble in anhydrous CH 2 Cl 2 (5 mL), TFA (1 mL) was added dropwise to the reaction solution, and after stirring at room temperature for 3 hours, the reaction mixture was concentrated in vacuo. Dissolve ...

Embodiment 2

[0067] Preparation of fluorescent probe Ⅰ-2

[0068] The preparation method is the same as in Example 1, with (7-((2-(3-azidopropoxy)-5-chlorophenyl)amino)heptyl)carbamate tert-butyl ester (0.878g, 2.0mmol) replacing (3-((2-(2-azidoethoxy)-5-fluorophenyl)amino)propyl)carbamate tert-butyl ester, 0.524g of yellow solid was obtained, the three-step yield was 27.6%, R f : 0.11 (methanol:ethyl acetate=2:5), m.p.159.7-160.1°C, the compound 1 H NMR (500MHz, DMSO-d 6 )δ9.10(s,1H),8.87(d,J=1.3Hz,1H),8.34(s,1H),8.20(s,1H),7.75–7.69(m,2H),7.65(d,J =2.0Hz,1H),7.27–7.19(m,2H),7.14(dd,J=10.2,7.4Hz,2H),7.01(dd,J=7.5,2.0Hz,1H),7.00(s,2H) ,6.78–6.70(m,2H),6.64–6.57(m,2H),6.50(d,J=2.1Hz,1H),6.34(dd,J=7.5,2.0Hz,1H),4.55(td,J =12.4,1.9Hz,1H),4.40(s,1H),4.23(d,J=12.3Hz,1H),4.14(d,J=12.4Hz,1H),3.91(ddt,J=20.2,12.2, 2.9Hz, 2H), 3.69–3.57(m, 1H), 3.40(dd, J=12.4, 10.9Hz, 1H), 3.28(td, J=12.0, 1.2Hz, 1H), 2.90(td, J=12.2 ,4.2Hz,1H),2.29–2.18(m,1H),2.04–1.54(m,6H),1.53–1.43(m,2H),1.44–1.25(m,2H)....

Embodiment 3

[0070] Preparation of fluorescent probe Ⅰ-3

[0071] The preparation method is the same as in Example 1, with (4-(2-(2-azidoethoxy)-5-iodophenoxy) butyl) tert-butyl carbamate (0.950g, 2.0mmol) replacing (3 -((2-(2-azidoethoxy)-5-fluorophenyl)amino)propyl)carbamate tert-butyl to give yellow solid 0.586g, three-step yield 29.8%, R f : 0.19 (methanol:ethyl acetate=1:2), m.p.161.7-162.8°C, the compound 1 H NMR (500MHz, DMSO-d 6 )δ9.59(s,1H),8.99(d,J=1.3Hz,1H),8.57(s,1H),8.20(s,1H),7.72(dd,J=7.5,2.0Hz,1H), 7.68–7.61(m,3H),7.42–7.34(m,2H),7.30(s,1H),7.16–7.10(m,1H),7.00(s,2H),6.85(dd,J=19.8,7.5 Hz,2H),6.52(dd,J=7.4,1.9Hz,1H),6.38(s,1H),5.09–4.96(m,2H),4.79–4.65(m,2H),4.58(ddt,J= 12.5,4.8,1.4Hz,1H), 4.36(ddd,J=12.6,11.4,4.0Hz,1H),4.17–4.08(m,2H),4.01–3.89(m,2H),2.32(qt,J= 12.4,3.3Hz,1H),2.21–2.10(m,1H),2.06–1.95(m,1H),1.81(dd,J=12.2,8.6,Hz,1H)., 13 C NMR (125MHz, DMSO-d 6 )δ179.64,172.72,170.00,168.45,158.76,153.94,151.93,150.97,149.24,146.85,146.41,142.66,139.43,137.31,136.03...

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Abstract

The invention relates to the field of chemical biology, and particularly, relates to small molecular fluorescent probes (I) designed based on prolyl hydroxylase inhibitors. The structural modules of the probes include prolyl hydroxylase binding groups, connection chains and fluorescent reporting groups. The invention also discloses a preparation method of the probes. The prolyl hydroxylase small molecular fluorescent probes can be used for high throughput screening of the prolyl hydroxylase inhibitors, and guide the discovery and structural optimization of the prolyl hydroxylase inhibitors. Astool molecules, the probes with the structures can quickly and accurately confirm the binding condition of small molecules with target proteins.

Description

technical field [0001] The present invention relates to the field of chemical biology. It specifically relates to a class of small molecule fluorescent probes designed based on prolyl hydroxylase inhibitors, its preparation method, and its application in determining the inhibitory activity of compounds on prolyl hydroxylase, which can be used for prolyl hydroxylation High-throughput screening of enzyme inhibitors can guide the discovery and structure optimization of prolyl hydroxylase inhibitors, and is suitable for guiding the discovery and treatment of anemia, ischemic disease and kidney injury, etc., which are closely related to prolyl hydroxylase Small molecule drugs for related diseases. Background technique [0002] Renal anemia, as one of the most common complications of chronic kidney disease patients, seriously reduces the quality of life of patients with chronic kidney disease, and also leads to increased morbidity and mortality of cardiovascular diseases. The ma...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D493/10C09K11/06G01N21/64
CPCC07D493/10C09K11/06G01N21/6428G01N21/6445
Inventor 尤启冬张晓进李治红
Owner CHINA PHARM UNIV
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