Identification Of Small Molecule Inhibitors Based On DYRK1A Protein Structure And Evaluation Of Their Activity

Overexpressed dual-specificity tyrosine-(Y)-phosphorylation-regulated kinase 1A(DYRK1A)is abnormally activated in pancreatic cancer and diabete,which plays an imortant role in tumorigenesis and the proliferation of islet β cells.Therefore,inhibition of DYRK1A might represent a novel approach,leading to more effective anti-cancer and diabetes therapies.A large number of high-purity recombinant human DYRK1A proteins were obtained through genetic engineering technology.DYRK1A gene was constructed on the new expres-sion vector pNlC28-Bsa4 by recombinant DNA technology,which realized the stable exis-tence and efficient cloning of DYRK1A gene.Here we discoved licocoumarone(LC)and 2-2 as potential DYRK1A inhibitors,with IC50 values of 12.56 μM and 3.44 μM,respectively.The molecular docking indicated these compounds completely occupied the whole pocket of DYRK1A and predicted the binding pattern between compounds and DYRK1A.The MST result indicated that LC had strong binding affinity to DYRK1A with the Kd value of 14.90μM.DARTS showed that LC could increase the stability of DYRK1A protein,indicated that DYRK1A was a potential target for LC.Meanwhile,LC induced significant cytotoxicity and apoptosis against DYRK1 A-overexpressing BxPC-3 cells.Treatment of BxPC-3 cells with LC obviously reduced c-MET protein expression level.According to the chemical structure of compound DMB,we obtained compounds 2-1 and 2-2 by chemical synthesis.Combining the in vitro DYRK1A kinase activity and MST assays,the structure-activity relationship of DYRK1A inhibitors was briefly analyzed.The results showed that hydroxyl group was likely to be the key pharmacophore;hydrogen bonds and hydrophobic interactions played an impor-tant role in the stability of the composite structure.This study established a mul-ti-component-single target screening model for the discovery of DYRK1A small molecule in-hibitors.Collectively,we organically combined molecular docking with the natural product data-base,combined protein biochemistry,pharmacological experiments and chemical synthesis,which obtained lead compounds as DYRK1A inhibitor for treating major disease.Licocou-marone and 2-2 were first identified as natural DYRK1A inhibitors,and they will play a lead-ing role in the discovery of drugs for treating human diseases,especially pancreatic cancer and diabetes mellitus.