ABSTRACT
The dual-specificity tyrosine-regulated kinases DYRK1A and DYRK1B play a key role in controlling the quiescence-proliferation switch in cancer cells. Serum reduction of U87MG 2D cultures or multi-cellular tumour spheroids induced a quiescent like state characterized by increased DYRK1B and p27, and decreased pRb and cyclin D1. VER-239353 is a potent, selective inhibitor of the DYRK1A and DYRK1B kinases identified through fragment and structure-guided drug discovery. Inhibition of DYRK1A/B by VER-239353 in quiescent U87MG cells increased pRb, DYRK1B and cyclin D1 but also increased the cell cycle inhibitors p21 and p27. This resulted in exit from G0 but subsequent arrest in G1. DYRK1A/B inhibition reduced the proliferation of U87MG cells in 2D and 3D culture with greater effects observed under reduced serum conditions. Paradoxically, the induced re-expression of cell cycle proteins by DYRK1A/B inhibition further inhibited cell proliferation. Cell growth arrest induced in quiescent cells by DYRK1A/B inhibition was reversible through the addition of growth-promoting factors. DYRK inhibition-induced DNA damage and synergized with a CHK1 inhibitor in the U87MG spheroids. In vivo, DYRK1A/B inhibition-induced tumour stasis in a U87MG tumour xenograft model. These results suggest that further evaluation of VER-239353 as a treatment for glioblastoma is therefore warranted.
Subject(s)
Cyclin D1/metabolism , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Cyclin-Dependent Kinase Inhibitor p27/metabolism , Glioblastoma/metabolism , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/antagonists & inhibitors , Signal Transduction/drug effects , Animals , Cell Cycle/drug effects , Cell Line, Tumor , Disease Models, Animal , Female , Glioblastoma/drug therapy , Glioblastoma/etiology , Glioblastoma/pathology , Humans , Mice , Protein Kinase Inhibitors/pharmacology , Dyrk KinasesABSTRACT
The kinase DYRK1A is an attractive target for drug discovery programs due to its implication in multiple diseases. Through a fragment screen, we identified a simple biaryl compound that is bound to the DYRK1A ATP site with very high efficiency, although with limited selectivity. Structure-guided optimization cycles enabled us to convert this fragment hit into potent and selective DYRK1A inhibitors. Exploiting the structural differences in DYRK1A and its close homologue DYRK2, we were able to fine-tune the selectivity of our inhibitors. Our best compounds potently inhibited DYRK1A in the cell culture and in vivo and demonstrated drug-like properties. The inhibition of DYRK1A in vivo translated into dose-dependent tumor growth inhibition in a model of ovarian carcinoma.
Subject(s)
Drug Design , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/antagonists & inhibitors , Adenosine Triphosphate/chemistry , Animals , Binding Sites , Cell Line, Tumor , Cell Survival/drug effects , Cyclin-Dependent Kinase 9/antagonists & inhibitors , Cyclin-Dependent Kinase 9/metabolism , Drug Evaluation, Preclinical , Female , Humans , Mice , Mice, Nude , Molecular Docking Simulation , Ovarian Neoplasms/drug therapy , Ovarian Neoplasms/pathology , Phosphorylation/drug effects , Protein Isoforms/antagonists & inhibitors , Protein Isoforms/metabolism , Protein Kinase Inhibitors/metabolism , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Protein Serine-Threonine Kinases/metabolism , Protein-Tyrosine Kinases/metabolism , Pyrimidines/chemistry , Pyrimidines/metabolism , Pyrimidines/pharmacology , Pyrimidines/therapeutic use , Structure-Activity Relationship , Dyrk KinasesABSTRACT
UDP-3-O-acyl-N-acetylglucosamine deacetylase (LpxC) is a zinc metalloenzyme that catalyzes the first committed step in the biosynthesis of Lipid A, an essential component of the cell envelope of Gram-negative bacteria. The most advanced, disclosed LpxC inhibitors showing antibacterial activity coordinate zinc through a hydroxamate moiety with concerns about binding to other metalloenzymes. Here, we describe the discovery, optimization, and efficacy of two series of compounds derived from fragments with differing modes of zinc chelation. A series was evolved from a fragment where a glycine moiety complexes zinc, which achieved low nanomolar potency in an enzyme functional assay but poor antibacterial activity on cell cultures. A second series was based on a fragment that chelated zinc through an imidazole moiety. Structure-guided design led to a 2-(1S-hydroxyethyl)-imidazole derivative exhibiting low nanomolar inhibition of LpxC and a minimum inhibitory concentration (MIC) of 4 µg/mL against Pseudomonas aeruginosa, which is little affected by the presence of albumin.
