ABSTRACT
The discovery of a highly potent and selective small molecule inhibitor 9 for in vitro target validation of MNK1/2 kinases is described. The aminopyrazine benzimidazole series was derived from an HTS hit and optimized by utilization of a docking model, conformation analysis, and binding pocket comparison against antitargets.
Subject(s)
Intracellular Signaling Peptides and Proteins/antagonists & inhibitors , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Binding Sites , Caco-2 Cells/drug effects , Drug Design , High-Throughput Screening Assays/methods , Humans , Intracellular Signaling Peptides and Proteins/chemistry , Intracellular Signaling Peptides and Proteins/metabolism , Molecular Docking Simulation , Permeability , Protein Serine-Threonine Kinases/chemistry , Protein Serine-Threonine Kinases/metabolism , Solubility , Structure-Activity RelationshipABSTRACT
BACKGROUND: Protein synthesis is tightly regulated and alterations to translation are characteristic of many cancers.Translation regulation is largely exerted at initiation through the eukaryotic translation initiation factor 4 F (eIF4F). eIF4F is pivotal for oncogenic signaling as it integrates mitogenic signals to amplify production of pro-growth and pro-survival factors. Convergence of these signals on eIF4F positions this factor as a gatekeeper of malignant fate. While the oncogenic properties of eIF4F have been characterized, genome-wide evaluation of eIF4F translational output is incomplete yet critical for developing novel translation-targeted therapies. RESULTS: To understand the impact of eIF4F on malignancy, we utilized a genome-wide ribosome profiling approach to identify eIF4F-driven mRNAs in MDA-MB-231 breast cancer cells. Using Silvestrol, a selective eIF4A inhibitor, we identify 284 genes that rely on eIF4A for efficient translation. Our screen confirmed several known eIF4F-dependent genes and identified many unrecognized targets of translation regulation. We show that 5'UTR complexity determines Silvestrol-sensitivity and altering 5'UTR structure modifies translational output. We highlight physiological implications of eIF4A inhibition, providing mechanistic insight into eIF4F pro-oncogenic activity. CONCLUSIONS: Here we describe the transcriptome-wide consequence of eIF4A inhibition in malignant cells, define mRNA features that confer eIF4A dependence, and provide genetic support for Silvestrol's anti-oncogenic properties. Importantly, our results show that eIF4A inhibition alters translation of an mRNA subset distinct from those affected by mTOR-mediated eIF4E inhibition. These results have significant implications for therapeutically targeting translation and underscore a dynamic role for eIF4F in remodeling the proteome toward malignancy.
Subject(s)
DEAD-box RNA Helicases/physiology , Eukaryotic Initiation Factor-4A/physiology , Transcriptome/drug effects , Apoptosis/drug effects , Apoptosis/genetics , Cell Cycle Checkpoints/drug effects , Cell Cycle Checkpoints/genetics , Cell Line, Tumor , DEAD-box RNA Helicases/antagonists & inhibitors , DEAD-box RNA Helicases/metabolism , Eukaryotic Initiation Factor-4A/antagonists & inhibitors , Eukaryotic Initiation Factor-4A/metabolism , Gene Expression Regulation, Neoplastic , Humans , Protein Biosynthesis/drug effects , Protein Biosynthesis/physiology , RNA, Messenger/metabolism , Triterpenes/pharmacologyABSTRACT
Numerous lines of evidence suggest that the polypeptide hormone prolactin (PRL) may contribute to breast and prostate tumorigenesis through its interactions with the prolactin receptor (PRLR). Here, we describe the biologic properties of LFA102, a humanized neutralizing monoclonal antibody directed against the extracellular domain of PRLR. This antibody was found to effectively antagonize PRL-induced signaling in breast cancer cells in vitro and in vivo and to block PRL-induced proliferation in numerous cell line models, including examples of autocrine/paracrine PRL activity. A single administration of LFA102 resulted in regression of PRL-dependent Nb2-11 tumor xenografts and significantly prolonged time to progression. Finally, LFA102 treatment significantly inhibited PRLR signaling as well as tumor growth in a carcinogen-induced, estrogen receptor-positive rat mammary cancer model as a monotherapy and enhanced the efficacy of the aromatase inhibitor letrozole when administered in combination. The biologic properties of LFA102, elucidated by the preclinical studies presented here, suggest that this antibody has the potential to be a first-in-class, effective therapeutic for the treatment of PRL-dependent cancers.