ABSTRACT
The prenyl-binding protein PDEδ is crucial for the plasma membrane localization of prenylated Ras. Recently, we have reported that the small-molecule Deltarasin binds to the prenyl-binding pocket of PDEδ, and impairs Ras enrichment at the plasma membrane, thereby affecting the proliferation of KRas-dependent human pancreatic ductal adenocarcinoma cell lines. Here, using structure-based compound design, we have now identified pyrazolopyridazinones as a novel, unrelated chemotype that binds to the prenyl-binding pocket of PDEδ with high affinity, thereby displacing prenylated Ras proteins in cells. Our results show that the new PDEδ inhibitor, named Deltazinone 1, is highly selective, exhibits less unspecific cytotoxicity than the previously reported Deltarasin and demonstrates a high correlation with the phenotypic effect of PDEδ knockdown in a set of human pancreatic cancer cell lines.
Subject(s)
Antineoplastic Agents/chemistry , Cyclic Nucleotide Phosphodiesterases, Type 6/chemistry , Epithelial Cells/drug effects , Gene Expression Regulation, Neoplastic , Phosphodiesterase Inhibitors/chemistry , Proto-Oncogene Proteins p21(ras)/chemistry , Pyrazines/chemistry , Pyrazoles/chemistry , Small Molecule Libraries/chemistry , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Benzimidazoles/chemistry , Benzimidazoles/pharmacology , Binding Sites , Cell Line, Tumor , Crystallography, X-Ray , Cyclic Nucleotide Phosphodiesterases, Type 6/antagonists & inhibitors , Cyclic Nucleotide Phosphodiesterases, Type 6/genetics , Cyclic Nucleotide Phosphodiesterases, Type 6/metabolism , Epithelial Cells/metabolism , Epithelial Cells/pathology , Gene Expression , Humans , Molecular Docking Simulation , Pancreatic Ducts/drug effects , Pancreatic Ducts/metabolism , Pancreatic Ducts/pathology , Phosphodiesterase Inhibitors/chemical synthesis , Phosphodiesterase Inhibitors/pharmacology , Protein Binding , Protein Interaction Domains and Motifs , Protein Structure, Secondary , Proto-Oncogene Proteins p21(ras)/antagonists & inhibitors , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Pyrazines/chemical synthesis , Pyrazines/pharmacology , Pyrazoles/chemical synthesis , Pyrazoles/pharmacology , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Signal Transduction , Small Molecule Libraries/chemical synthesis , Small Molecule Libraries/pharmacologyABSTRACT
The X-linked dominant trait focal dermal hypoplasia (FDH, Goltz syndrome) is a developmental defect with focal distribution of affected tissues due to a block of Wnt signal transmission from cells carrying a detrimental PORCN mutation on an active X-chromosome. Molecular characterization of 24 unrelated patients from different ethnic backgrounds revealed 23 different mutations of the PORCN gene in Xp11.23. Three were microdeletions eliminating PORCN and encompassing neighboring genes such as EBP, the gene associated with Conradi-Hünermann-Happle syndrome (CDPX2). 12/24 patients carried nonsense mutations resulting in loss of function. In one case a canonical splice acceptor site was mutated, and 8 missense mutations exchanged highly conserved amino acids. FDH patients overcome the consequences of potentially lethal X-chromosomal mutations by extreme skewing of X-chromosome inactivation in females, enabling transmission of the trait in families, or by postzygotic mosaicism both in male and female individuals. Molecular characterization of the PORCN mutations in cases diagnosed as Goltz syndrome is particularly relevant for genetic counseling of patients and their families since no functional diagnostic test is available and carriers of the mutation might otherwise be overlooked due to considerable phenotypic variability associated with the mosaic status.
