生物活性

AZ191 is a potent and selective DYRK1B inhibitor with IC50 of 17 nM in a cell-free assay, about 5- and 110-fold selectivity over DYRK1A and DYRK2, respectively. Inhibits DYRK1B-induced Gi phase cell cycle arrest in Panc-1 cells in vitro.

体外研究

体内研究

激酶实验

In vitro DYRK kinase assay[1]

EGFP–DYRK1A, EGFP–DYRK1B or EGFP–DYRK2 was immunoprecipitatedfrom whole-cell lysate and pre-incubated in the absence or presence of variousconcentrations of AZ191 or harmine for 5min at room temperature (20 ℃) and thenassayed for kinase activity by incubating with 50μM Woodtide, with two additionallysine residues attached to the N-terminus to allow it to bind to P81 paper(KKSSCYVDRKIYTYIQSRFY), 50mM Tris/HCl (pH 7.5), 0.1mM EGTA, 0.1%2-mercaptoethanol, 10 mM MgCl2 and 0.1mM [γ -³²P]ATP in a totalvolume of 50μl for 25 min at 30 ^◦C.

细胞实验

Cells and cell culture[1]

HEK (human embryonic kidney)-293 and PANC-1cells were maintained in DMEM (Dulbecco’s modified Eagle’s medium) supplementedwith 10% FBS, 2mM L-glutamine, 100 units/ml penicillin and 0.1μg/mlstreptomycin. HTetR cells were maintained in DMEM supplemented with 10% FBS,2mM L-glutamine, 100 units/ml penicillin, 0.1μg/ml streptomycin and 5μg/mlblasticidin to maintain stable clone selection. HD1B cells were maintained inDMEM supplemented with 10%FBS, 2mM L-glutamine, 100 units/ml penicillin,0.1μg/ml streptomycin, 5μg/ml blasticidin and 100μg/ml ZeocinTM. Cells wereroutinely passaged until they reached 80% confluency.

Preparationof cell extracts and Western blotting

Cells were lysed in ice-cold Tris-glycinelysis buffer, assayed for protein content and fractionated by SDS/PAGE. TheSDS/PAGE gels were transferred on to Immobilon P membranes, which were blockedin 0.1% Tween 20/TBS containing 5% (w/v) powdered milk and probed with theindicated antibodies. Immune complexes were visualized with the ECL system.

动物实验

不同实验动物依据体表面积的等效剂量转换表（数据来源于FDA指南）

例如，已知某工具药用于小鼠的剂量为88 mg/kg , 则用于大鼠的剂量换算方法：将88 mg/kg 乘以小鼠的Km系数（3），再除以大鼠的Km系数（6），得到该药物用于大鼠的等效剂量44 mg/kg。

参考文献

[1] Ashford AL, Oxley D, Kettle J, et al. A novel DYRK1B inhibitor AZ191 demonstrates that DYRK1B acts independently of GSK3beta to phosphorylate cyclin D1 at Thr(286), not Thr(288). Biochem J. 2014;457(1):43-56.

体内溶解度

Clinical Trial Information ( data from http://clinicaltrials.gov )

注：以上所有数据均来自公开文献，并不保证对所有实验均有效，数据仅供参考。

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