RESUMEN
BACKGROUND: The myotonic dystrophy kinase-related CDC42-binding kinases MRCKα and MRCKß regulate actin-myosin contractility and have been implicated in cancer metastasis. Along with the related ROCK1 and ROCK2 kinases, the MRCK proteins initiate signalling events that lead to contractile force generation which powers cancer cell motility and invasion. A potential strategy for cancer therapy is to reduce metastasis by blocking MRCK activity, either alone or in combination with ROCK inhibition. However, to date no potent small molecule inhibitors have been developed with selectivity towards MRCK. RESULTS: Screening a kinase-focused small molecule chemical library resulted in the identification of compounds with inhibitory activity towards MRCK. Medicinal chemistry combined with in vitro enzyme profiling led to the discovery of 4-chloro-1-(4-piperidyl)-N-[5-(2-pyridyl)-1H-pyrazol-4-yl]pyrazole-3-carboxamide (BDP00005290; abbreviated as BDP5290) as a potent MRCK inhibitor. X-ray crystallography of the MRCKß kinase domain in complex with BDP5290 revealed how this ligand interacts with the nucleotide binding pocket. BDP5290 demonstrated marked selectivity for MRCKß over ROCK1 or ROCK2 for inhibition of myosin II light chain (MLC) phosphorylation in cells. While BDP5290 was able to block MLC phosphorylation at both cytoplasmic actin stress fibres and peripheral cortical actin bundles, the ROCK selective inhibitor Y27632 primarily reduced MLC phosphorylation on stress fibres. BDP5290 was also more effective at reducing MDA-MB-231 breast cancer cell invasion through Matrigel than Y27632. Finally, the ability of human SCC12 squamous cell carcinoma cells to invade a three-dimensional collagen matrix was strongly inhibited by 2 µM BDP5290 but not the identical concentration of Y27632, despite equivalent inhibition of MLC phosphorylation. CONCLUSIONS: BDP5290 is a potent MRCK inhibitor with activity in cells, resulting in reduced MLC phosphorylation, cell motility and tumour cell invasion. The discovery of this compound will enable further investigations into the biological activities of MRCK proteins and their contributions to cancer progression.
Asunto(s)
Antineoplásicos/farmacología , Proteína Quinasa de Distrofia Miotónica/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles/farmacología , Piridinas/farmacología , Amidas/farmacología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Humanos , Proteína Quinasa de Distrofia Miotónica/metabolismo , Invasividad Neoplásica , Quinasas Asociadas a rho/antagonistas & inhibidores , Quinasas Asociadas a rho/metabolismoRESUMEN
Based on the theoretical understanding of the in vivo lysosomotropism, by adjusting the pk(a) of basic nitrogen containing cathepsin S inhibitors, a set of compounds with pk(a) 6-8 were identified to have excellent cell based Lip10 activity, yet avoiding undesired sequestration in spleen.
Asunto(s)
Catepsinas/antagonistas & inhibidores , Nitrógeno/química , Inhibidores de Proteasas/química , Piridinas/química , Animales , Catepsinas/metabolismo , Ratones , Inhibidores de Proteasas/síntesis química , Inhibidores de Proteasas/farmacocinética , Piridinas/síntesis química , Piridinas/farmacocinética , Ratas , Ratas Sprague-DawleyRESUMEN
The trifluoromethylphenyl P2 motif from previously reported heteroarylnitrile series has been successfully applied for the design and synthesis of highly potent novel ketoamide-based cathepsin S inhibitors. The key in this process is the change of the torsion angle between the P2 phenyl ring and the attached secondary amide by adding a small Cl, F, or Me group at the 2-position.
Asunto(s)
Compuestos de Anilina/síntesis química , Catepsinas/antagonistas & inhibidores , Inhibidores de Cisteína Proteinasa/síntesis química , Amidas/síntesis química , Amidas/farmacología , Compuestos de Anilina/farmacología , Animales , Inhibidores de Cisteína Proteinasa/farmacología , Flúor , Humanos , Cetonas , Relación Estructura-ActividadRESUMEN
The myotonic dystrophy-related Cdc42-binding kinases MRCKα and MRCKß contribute to the regulation of actin-myosin cytoskeleton organization and dynamics, acting in concert with the Rho-associated coiled-coil kinases ROCK1 and ROCK2. The absence of highly potent and selective MRCK inhibitors has resulted in relatively little knowledge of the potential roles of these kinases in cancer. Here, we report the discovery of the azaindole compounds BDP8900 and BDP9066 as potent and selective MRCK inhibitors that reduce substrate phosphorylation, leading to morphologic changes in cancer cells along with inhibition of their motility and invasive character. In over 750 human cancer cell lines tested, BDP8900 and BDP9066 displayed consistent antiproliferative effects with greatest activity in hematologic cancer cells. Mass spectrometry identified MRCKα S1003 as an autophosphorylation site, enabling development of a phosphorylation-sensitive antibody tool to report on MRCKα status in tumor specimens. In a two-stage chemical carcinogenesis model of murine squamous cell carcinoma, topical treatments reduced MRCKα S1003 autophosphorylation and skin papilloma outgrowth. In parallel work, we validated a phospho-selective antibody with the capability to monitor drug pharmacodynamics. Taken together, our findings establish an important oncogenic role for MRCK in cancer, and they offer an initial preclinical proof of concept for MRCK inhibition as a valid therapeutic strategy.Significance: The development of selective small-molecule inhibitors of the Cdc42-binding MRCK kinases reveals their essential roles in cancer cell viability, migration, and invasive character. Cancer Res; 78(8); 2096-114. ©2018 AACR.
Asunto(s)
Antineoplásicos/uso terapéutico , Carcinoma de Células Escamosas/tratamiento farmacológico , Descubrimiento de Drogas , Proteína Quinasa de Distrofia Miotónica/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/uso terapéutico , Piridinas/uso terapéutico , Pirimidinas/uso terapéutico , Pirroles/uso terapéutico , Neoplasias Cutáneas/tratamiento farmacológico , Animales , Antineoplásicos/farmacología , Carcinoma de Células Escamosas/enzimología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Células HEK293 , Humanos , Ratones , Ratones Desnudos , Fosforilación , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/farmacología , Pirimidinas/farmacología , Pirroles/farmacología , Neoplasias Cutáneas/enzimología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We report the discovery of a benzimidazole series of CYP11B2 inhibitors. Hit-to-lead and lead optimization studies identified compounds such as 32, which displays potent CYP11B2 inhibition, high selectivity versus related CYP targets, and good pharmacokinetic properties in rat and rhesus. In a rhesus pharmacodynamic model, 32 produces dose-dependent aldosterone lowering efficacy, with no apparent effect on cortisol levels.