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Preparation method and application of BETHDAC dual-targeting PROTAC

A dual-target, unsaturated technology, applied in the field of medicine

Pending Publication Date: 2021-11-16
SHANGHAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Theoretically, using PROTACs to achieve double knockout of BRD4 and HDACs will overcome the problems of dual-target inhibitors, and there is no literature report on the simultaneous double knockout of two target proteins using rationally designed PROTACs

Method used

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  • Preparation method and application of BETHDAC dual-targeting PROTAC
  • Preparation method and application of BETHDAC dual-targeting PROTAC
  • Preparation method and application of BETHDAC dual-targeting PROTAC

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] 8- (2 - ((S) -4- (4- chlorophenyl) -2,3,9-trimethyl--6H- thieno [3,2-f] [1,2,4] triazole [4,3-a] [1,4] diazepin-1-yl) -N- (2- (2- (2- (2 - ((2- (2,6- dioxo-piperidin - 3- yl) 1-oxo-isoindol-4-yl) amino) -2-oxo-ethoxy) ethoxy) ethoxy) ethyl) acetamido) -N- hydroxyoctanoate amide synthesis of ( M10)

[0059] Step a: tert-butyl (2- (2- (2- (2- (2 - ((2- (2,6-dioxo-piperidin-3-yl) isoindole-1-oxo-4 - yl) amino) -2-oxo-ethoxy) ethoxy) ethoxy) synthesis of urethane (M2)

[0060] Step a: Compound M0 (1.0g, 3.1mmol) H, ATU (2.4g, 6.2mmol) D, IPEA (1.0mL, 7.8mmol) was dissolved in dry DMF, the reaction at room temperature for 0.5h and then added to lenalidomide M1 (0.80g, 3.1mmol) the reaction after 4h, the reaction was complete by TLC. The reaction mixture was slowly poured into ice water (1.5 L of) mixture with EA (30mL × 3) After extraction the organic phases were combined, solvent was distilled off under reduced pressure, the residue was purified by silica gel column chromatogra...

Embodiment 2

[0076] Example 2: inhibitory activity of the present compounds tested (Ki) to BRD4 and HDAC1.

[0077] 5μL The test compound (concentration of each dilution), BRD4 (20nM) and PMDM6-F (20nM) (Buffer: 100mM tripotassium phosphate, PH = 7.5; 100μg / mL BGG; 0.02% sodium azide) was added to 96-well black after the flat microtiter plates to a final volume of 115μL, incubated at room temperature for 1 hour, Biotek-Synergy plate reader (excitation light of 485 nM, the emitted light is 528 nm) value of fluorescence polarization readings.

[0078]The fluorescence polarization value obtained by the above method is to draw curves with Origin 9.0 software, and the protein binding inhibitory constant (ki) is calculated. HDAC1 test method is BRD4.

[0079] Experimental results: First test the inhibitory activity of all target compounds to BRD4 and HDAC1 proteins, and select (+) - JQ1 and Saha as a positive control. The test results are shown in Table 4-2, and the compound M10 retaining the BRD4...

Embodiment 3

[0082] Example 3: In vitro anti-tumor activity test of the compound of the present invention (IC 50 .

[0083] The compounds of the present invention have been tested by three tumor cell proliferation and suppression capability testing, and the test method adopts a conventional CCK8 method. Tumor cells (MCF-7 (human pulmonary cancer cells), A549 (human lung cancer cells), HepG2 (human lung cancer cells), HepG2 (human liver cancer cells), HepG2 (human lung cancer cells), then medium (DMEM + 10% FBS or Prmi1640) + 10% FBS) Dilute the cells suspended into a single cell suspension, adjust the cell density of 5 × 10 4 Add 100 μl of inoculated in 96-well plates per well, 37 ° C, 5% CO 2 In the incubator for 24 hours, different concentrations of compounds were added, and each concentration was three complexes, and the experimental group and control group were set, and after 72 hours of incubation, 10 μl Ck8 solution was added to each well, and then 37 ° C After 1-4 hours of light, the 45...

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PUM

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Abstract

The invention discloses a preparation method and application of BETHDAC dual-targeting PROTAC. The compound has a structure as shown in a general formula (I). The compound is reasonably designed through a BET inhibitor ((+)-JQ1), an HDAC inhibitor and E3 ubiquitin ligase. Pharmacological experiments show that the compound has relatively strong binding inhibition activity on BET protein and HDAC protein and in-vitro anti-tumor proliferation activity. Mechanism experiments show that the compound BET / HDAC double-target PROTAC can obviously induce degradation of the BRD4 protein and the HDACs protein, and can be applied to tumor diseases with pathological characteristics mediated by the BRD4 protein and the HDACs protein. The compound as a BET / HDAC double-target PROTAC molecule reported for the first time has further development and research values.

Description

Technical field [0001] The invention relates to the field of medical technology, specifically a preparation method and application of BETHDAC dual-targeting PROTAC, and the application of this type of small molecule compound in the treatment of malignant tumor diseases. Background technique [0002] As an emerging technology, PROTAC uses bifunctional small molecules to make the target protein recognized by E3 ubiquitin ligase and labeled with ubiquitination, and then uses the cell's own ubiquitin-proteasome system to degrade the target protein. Currently, most research on PROTACs focuses on degrading a single target protein. Although some literature has been published that down-regulation of two or more proteins can be achieved, most of them are caused by PROTAC off-target effects, or are the result of inhibiting upstream signals causing down-regulation of multiple downstream proteins, or due to acting on The ligand pockets of the same type of protein result, so PROTAC mole...

Claims

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Application Information

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IPC IPC(8): C07D495/14A61P35/00
CPCC07D495/14A61P35/00
Inventor 何世鹏马俊辉余自强刘莹马浩骞陈宝宝纪雅静
Owner SHANGHAI UNIV
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