ABSTRACT
The c-Jun N-terminal kinases (JNK-1, -2, and -3) are members of the mitogen activated protein (MAP) kinase family of enzymes. They are activated in response to certain cytokines, as well as by cellular stresses including chemotoxins, peroxides, and irradiation. They have been implicated in the pathology of a variety of different diseases with an inflammatory component including asthma, stroke, Alzheimer's disease, and type 2 diabetes mellitus. In this work, high-throughput screening identified a JNK inhibitor with an excellent kinase selectivity profile. Using X-ray crystallography and biochemical screening to guide our lead optimization, we prepared compounds with inhibitory potencies in the low-double-digit nanomolar range, activity in whole cells, and pharmacokinetics suitable for in vivo use. The new compounds were over 1,000-fold selective for JNK-1 and -2 over other MAP kinases including ERK2, p38alpha, and p38delta and showed little inhibitory activity against a panel of 74 kinases.
Subject(s)
Aminopyridines/chemical synthesis , Mitogen-Activated Protein Kinase 8/antagonists & inhibitors , Mitogen-Activated Protein Kinase 9/antagonists & inhibitors , Aminopyridines/chemistry , Aminopyridines/pharmacology , Animals , Biological Availability , Cell Line, Tumor , Crystallography, X-Ray , Half-Life , Humans , Mitogen-Activated Protein Kinase 10/metabolism , Mitogen-Activated Protein Kinase 8/chemistry , Mitogen-Activated Protein Kinase 8/metabolism , Mitogen-Activated Protein Kinase 9/metabolism , Models, Molecular , Phosphorylation , Protein Conformation , Rats , Rats, Sprague-DawleyABSTRACT
Glucocorticoids are used clinically to treat a variety of inflammatory diseases including endotoxemia. We hypothesized that injecting mice with the steroid prednisolone (pred) would mitigate the enhanced bone marrow (BM) natural suppressor (NS) cell activity that occurs in mice after receiving an injection of lipopolysaccharide (LPS). In vitro, prednisolone blocked the ability of NS cells to produce the immunosuppressive molecule nitric oxide (NO) and also the ability to suppress T cell proliferation. Prednisolone acted both indirectly, by blocking synthesis of cytokines necessary for NS cell activation, and also directly on NS cells, by blocking production of NO. In vivo, variable results were obtained. Prednisolone at 20 microg/gm did decrease NS activity when injected into normal mice. However, when mice were injected with both LPS and prednisolone (0.2 or 20 microg/gm), a large increase in BM NS activity was observed. The increase was evident in both the ability of the BM cells to suppress T cell proliferation and to produce NO. The data show that, in vivo, the steroid prednisolone in conjunction with the inflammatory compound LPS act to enhance BM NS activity.
