title: A yeast-based chemical screen identifies a PDE inhibitor that elevates steroidogenesis in mouse Leydig cells via PDE8 and PDE4 inhibition.
abstract: A cell-based high-throughput screen (HTS) was developed to detect phosphodiesterase 8 (PDE8) and PDE4/8 combination inhibitors. By replacing the Schizosaccharomyces pombe PDE gene with the murine PDE8A1 gene in strains lacking adenylyl cyclase, we generated strains whose protein kinase A (PKA)-stimulated growth in 5-fluoro orotic acid (5FOA) medium reflects PDE8 activity. From our previously-identified PDE4 and PDE7 inhibitors, we identified a PDE4/8 inhibitor that allowed us to optimize screening conditions. Of 222,711 compounds screened, ∼0.2% displayed composite Z scores of >20. Additional yeast-based assays using the most effective 367 compounds identified 30 candidates for further characterization. Among these, compound BC8-15 displayed the lowest IC₅₀ value for both PDE4 and PDE8 inhibition in in vitro enzyme assays. This compound also displays significant activity against PDE10A and PDE11A. BC8-15 elevates steroidogenesis in mouse Leydig cells as a single pharmacological agent. Assays using BC8-15 and two structural derivatives support a model in which PDE8 is a primary regulator of testosterone production by Leydig cells, with an additional role for PDE4 in this process. BC8-15, BC8-15A, and BC8-15C, which are commercially available compounds, display distinct patterns of activity against PDE4, PDE8, PDE10A, and PDE11A, representing a chemical toolkit that could be used to examine the biological roles of these enzymes in cell culture systems.
authors: Biology Department, Boston College, Chestnut Hill, Massachusetts, United States of America.
authors: Didem
authors: D
authors: Demirbas
authors: Arlene R
authors: AR
authors: Wyman
authors: Masami
authors: M
authors: Shimizu-Albergine
authors: Ozgur
authors: O
authors: Cakici
authors: Joseph A
authors: JA
authors: Beavo
authors: Charles S
authors: CS
authors: Hoffman
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4 / Q000737:chemistry / Q000378:metabolism*; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D057166:High-Throughput Screening Assays; D006801:Humans; D007985:Leydig Cells / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D051379:Mice; D062105:Molecular Docking Simulation; D058988:Phosphodiesterase 4 Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D011487:Protein Conformation; D012568:Schizosaccharomyces / Q000235:genetics*; D054852:Small Molecule Libraries / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D013256:Steroids / Q000096:biosynthesis*
chemical_list: D058988:Phosphodiesterase 4 Inhibitors; D054852:Small Molecule Libraries; D013256:Steroids; D054703:Cyclic Nucleotide Phosphodiesterases, Type 4
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