论文部分内容阅读
AIM To further investigate the new binding mode and action of mechanism of a novel IKK-β inhibitor which can specifically interact with new site on IKK-β and then suppress inflammatory conditions in vitro and in vivo.METHODS In vitro and in vivo studies of identification of the anti-inflammatory action and mechanisms of Compound B derived from a medical plant were conducted.The biotinylated DMY (DMY-biotin) was synthesized as tool probe to confirm the direct interaction between DMY and IKK-β protein, their protein adduct was detected by western blotting using biotin specific streptavidin-HRP antibody.IKK-β displacement binding assay was used to confirm the novel drug binding site of DMY on IKK-β.RESULTS A novel drug binding site of IKKβ kinase at the Cys-46 residue was found, and the Cys-46 residue of IKK-β exhibited strong and unique capability of binding with small-molecule IKK-β inhibitor, Compound B derived from a medical plant, which potently suppressed in vitro and in vivo mouse inflammatory and rat arthritic conditions.Cys-179 residue and ATP binding site have been known as drug targeting sites for most of the existing IKK-β inhibitors, while the new Cys-46 residue of IKK-β is able to bind with Compound B and then overcome the in vitro drug resistant phenotype of IKK-β mutants with mutations on the Cys-179 residue and ATP binding site.Therefore, finding of Cys-46 could provide insights of the new biochemical properties of IKK-β kinase as well as its mechanisms of inhibition.This new finding may also provide possibility of establishing a new platform for screening new IKK-β inhibitors.Discovery of novel kinase inhibitors with different inhibitory mechanisms or with new drug binding property has been evident in highly demand to circumvent the drug-resistant phenotypes of mutant kinases acquired from somatic gene mutations towards highly personalized therapy of patients.Indeed, a number of gene mutations of IKBKB have been identified in human cancer genomes although with yet unknown pathological functions, possibly which will alter the pharmacological effects of the existing IKK-β inhibitors.Since in vitro study demonstrated that mutation of the ATP binding site and Cys-179 of IKK-β greatly affected the binding kinetics of the existing IKK-β inhibitors, and in clinic, patients developed drug resistance due to primary or acquired mutation is common.CONCLUSION It is believed that identification of new mode of drug actions and the binding sites of new IKK-β inhibitors like Compound B has significant potential for developing a variety of IKK-β inhibitors.At the same time, it may provide alternate treatment option for patients who might have chance to develop drug resistance with the currently available IKK-β inhibitors in future.