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A novel allosteric inhibitor that prevents IKK? activation.

ABSTRACT: I kappa B kinase ? (IKK?) is one of the primary targets to regulate canonical NF-?B activity. The misregulation of NF-?B is associated with various diseases, including chronic inflammation and cancers. Most of the known IKK? inhibitors target its active form and suffer from poor selectivity. In the present study, we aim to design inhibitors that can bind to the IKK? inactive form and block its activation. We identified a potential allosteric site between the kinase domain (KD) and ubiquitin-like domain (ULD) of human IKK? and used it to virtually screen a chemical library for allosteric inhibitors. Among the 133 compounds tested, 16 inhibited NF-?B activity by over 50% at 50 ?M in a reporter gene assay. Further quantitative measurements and cytotoxicity study gave one compound 124 (3,4-dichloro-2-ethoxy-N-(2,2,6,6-tetramethylpiperidin-4-yl)benzenesulfonamide) which specifically targets the IKK? inactive form. In cells, 124 inhibited I?B? phosphorylation and NF-?B transcriptional activity for the reporter gene with an IC50 of 35 ?M by decreasing the phosphorylation level of Ser177/181 on IKK? and blocking its activation upon TNF? stimulation. Molecular dynamics simulations demonstrated that 124 binds to the pocket between KD and ULD in the inactive conformation of IKK? rather than the active conformation. As the first allosteric inhibitor that prevents IKK? activation, 124 provides a good starting point for further inhibitor discovery and a probe for IKK? enzyme cycle and regulatory mechanism study.

SUBMITTER: Liu H

PROVIDER: S-EPMC6083782 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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A novel allosteric inhibitor that prevents IKKβ activation.

Liu Hongbo H Liang Hao H Meng Hu H Deng Xiaobing X Zhang Xiaoling X Lai Luhua L

MedChemComm 20180109 2

I kappa B kinase β (IKKβ) is one of the primary targets to regulate canonical NF-κB activity. The misregulation of NF-κB is associated with various diseases, including chronic inflammation and cancers. Most of the known IKKβ inhibitors target its active form and suffer from poor selectivity. In the present study, we aim to design inhibitors that can bind to the IKKβ inactive form and block its activation. We identified a potential allosteric site between the kinase domain (KD) and ubiquitin-like ...[more]

PMID: 30108917

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Project description:Essentials Signaling by Gas6 through Tyro3/Axl/Mer receptors is essential for stable platelet aggregation. UNC2025 is a small molecule inhibitor of the Mer tyrosine kinase. UNC2025 decreases platelet activation in vitro and thrombus formation in vivo. UNC2025's anti-platelet effect is synergistic with inhibition of the ADP receptor, P2Y12 . SUMMARY:Background Growth arrest-specific protein 6 signals through the TAM (TYRO-3-AXL-MERTK) receptor family, mediating platelet activation and thrombus formation via activation of the aggregate-stabilizing ?IIb ?3 integrin. Objective To describe the antithrombotic effects mediated by UNC2025, a small-molecule MERTK tyrosine kinase inhibitor. Methods MERTK phosphorylation and downstream signaling were assessed by immunoblotting. Light transmission aggregometry, flow cytometry and microfluidic analysis were used to evaluate the impact of MERTK inhibition on platelet activation and stability of aggregates in vitro. The effects of MERTK inhibition on arterial and venous thrombosis, platelet accumulation at microvascular injury sites and tail bleeding times were determined with murine models. The effects of combined treatment with ADP-P2Y1&12 pathway antagonists and UNC2025 were also evaluated. Results and Conclusions Treatment with UNC2025 inhibited MERTK phosphorylation and downstream activation of AKT and SRC, decreased platelet activation, and protected animals from pulmonary embolism and arterial thrombosis without increasing bleeding times. The antiplatelet effect of UNC2025 was enhanced in combination with ADP-P2Y1&12 pathway antagonists, and a greater than additive effect was observed when these two agents with different mechanisms of inhibition were coadministered. TAM kinase signaling represents a potential therapeutic target, as inhibition of this axis, especially in combination with ADP-P2Y pathway antagonism, mediates decreased platelet activation, aggregate stability, and thrombus formation, with less hemorrhagic potential than current treatment strategies. The data presented here also demonstrate antithrombotic activity mediated by UNC2025, a novel translational agent, and support the development of TAM kinase inhibitors for clinical applications.

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A novel allosteric inhibitor that prevents IKK? activation.

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A novel allosteric inhibitor that prevents IKKβ activation.

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