ABSTRACT
A challenge for biomedical research is the development of pharmaceuticals that appropriately target disease mechanisms. Natural products can be a rich source of bioactive chemicals for medicinal applications but can act through unknown mechanisms and can be difficult to produce or obtain. To address these challenges, we developed a new marine-derived, renewable natural products resource and a method for linking bioactive derivatives of this library to the proteins and biological processes that they target in cells. We used cell-based screening and computational analysis to match gene expression signatures produced by natural products to those produced by small interfering RNA (siRNA) and synthetic microRNA (miRNA) libraries. With this strategy, we matched proteins and miRNAs with diverse biological processes and also identified putative protein targets and mechanisms of action for several previously undescribed marine-derived natural products. We confirmed mechanistic relationships for selected siRNAs, miRNAs, and compounds with functional roles in autophagy, chemotaxis mediated by discoidin domain receptor 2, or activation of the kinase AKT. Thus, this approach may be an effective method for screening new drugs while simultaneously identifying their targets.
Subject(s)
Biological Products/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Gene Ontology , Transcriptome/drug effects , Animals , Autophagy/drug effects , Autophagy/genetics , Bacteria/chemistry , Bacteria/classification , Biological Products/chemistry , Biological Products/isolation & purification , Cell Line, Tumor , Cells, Cultured , Cluster Analysis , Computational Biology/methods , Drug Evaluation, Preclinical/methods , Gene Regulatory Networks/drug effects , Gene Regulatory Networks/genetics , HCT116 Cells , Humans , Invertebrates/chemistry , MCF-7 Cells , Marine Biology , Mice , Mice, Inbred BALB C , MicroRNAs/genetics , Molecular Structure , Oligonucleotide Array Sequence Analysis , RNA InterferenceABSTRACT
Discoidin domain receptor 2 (DDR2) is a receptor tyrosine kinase involved in a variety of cellular response pathways, including regulation of cell growth, proliferation, and motility. Using a newly developed platform to identify the signaling pathway/molecular target of natural products, we identified a family of alkaloid natural products, discoipyrroles A-D (1-4), from Bacillus hunanensis that inhibit the DDR2 signaling pathway. The structure of 1-4, determined by detailed two-dimensional (2D) NMR methods and confirmed by X-ray crystallographic analysis has an unusual 3H-benzo[d]pyrrolo][1,3]oxazine-3,5-dione core. Discoipyrroles A-D potently inhibit DDR2 dependent migration of BR5 fibroblasts and show selective cytotoxicity to DDR2 mutant lung cancer cell lines (IC50 120-400 nM). Examination of the biosynthesis has led to the conclusion that the discoipyrroles are formed through a nonenzymatic process, leading to a one-pot total synthesis of 1.
Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Bacillus/chemistry , Biological Products/pharmacology , Fibroblasts/drug effects , Heterocyclic Compounds, 3-Ring/pharmacology , Pyrrolidinones/pharmacology , Receptor Protein-Tyrosine Kinases/antagonists & inhibitors , Receptors, Mitogen/antagonists & inhibitors , Antineoplastic Agents, Phytogenic/chemistry , Antineoplastic Agents, Phytogenic/isolation & purification , Biological Products/chemistry , Biological Products/isolation & purification , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Cell Survival/drug effects , Discoidin Domain Receptors , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Fibroblasts/cytology , Heterocyclic Compounds, 3-Ring/chemistry , Heterocyclic Compounds, 3-Ring/isolation & purification , Humans , Models, Molecular , Molecular Structure , Pyrrolidinones/chemistry , Pyrrolidinones/isolation & purification , Receptor Protein-Tyrosine Kinases/genetics , Receptor Protein-Tyrosine Kinases/metabolism , Receptors, Mitogen/genetics , Receptors, Mitogen/metabolism , Signal Transduction/drug effects , Stereoisomerism , Structure-Activity RelationshipABSTRACT
RFamide neuropeptides NPFF and NPAF affect gene expression in mature 3T3-L1 adipocytes but their role on adipogenesis is unknown. Here, we show that NPFF, NPAF, and NPSF inhibited the differentiation of 3T3-F442A preadipocytes in a concentration-dependent manner, but had no effect on 3T3-L1 adipogenesis. All three neuropeptides also blocked the adipose differentiation of normal and lipoma-derived human preadipocytes. The antiadipogenic effect of RFamide neuropeptides was linked with the overexpression of Id3 gene and the inhibition by NPAF remained after neuropeptide removal and further incubation of 3T3 cells with adipogenic medium. Our results show that NPFF, NPAF and NPSF negatively affect adipogenesis and suggest that these compounds participate in the regulation of the adipose tissue development by the central nervous system.
