title: High-throughput screening of ecdysone agonists using a reporter gene assay followed by 3-D QSAR analysis of the molting hormonal activity.
abstract: In this study, 172 diacylhydrazine analogs were examined for their ability to activate an ecdysone (molting hormone)-dependent reporter gene in a silkworm (Bombyx mori) cell-based high-throughput screening assay. The measured EC(50) values (concentration required to cause an effect in 50% of the cells) were used to construct a 3-D QSAR model that describes the ecdysone agonist activities of the diacylhydrazine analogs. Of these compounds, 14 exhibited no activity and were excluded from the 3-D QSAR analysis. The resulting equation described approximately 74% of the activity for 158 compounds. The final equation consisted of 42% electrostatic and 58% steric effects (r(2) = 0.74 and q(2) = 0.45). Comparative molecular field analysis (CoMFA) was used to visualize the steric and electrostatic potential fields that were favorable and unfavorable for biological activity. Of particular interest was the observation that the hydrophobic parameter (logP) was not necessary for describing the observed activities, although previous studies have cited the importance of hydrophobic parameters in both classical and 3-D QSAR analyses of these compounds. Modeling studies of the B. mori ecdysone receptor supported the observed physicochemical parameters required for activity reported by the CoMFA models. Comparison of the present analysis with those performed using other lepidopteran assay systems evidenced a high degree of correlation (r(2) = 0.81 for a Sf-9 cell-based assay and r(2) = 0.89 for a Chilo suppressalis integument-based assay), indicating that it is valid to compare the results generated with the B. mori cell-based system to those generated with previous lepidopteran assays. This novel assay system is amendable to a high-throughput screening format and should greatly increase our ability to discover novel agonists of molting hormone (ecdysone) activity.
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Japan.
authors.forename: Craig E
authors.initials: CE
authors.lastname: Wheelock
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Nakagawa
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Harada
authors.forename: Nobuhiro
authors.initials: N
authors.lastname: Oikawa
authors.forename: Miki
authors.initials: M
authors.lastname: Akamatsu
authors.forename: Guy
authors.initials: G
authors.lastname: Smagghe
authors.forename: Dimitra
authors.initials: D
authors.lastname: Stefanou
authors.forename: Kostas
authors.initials: K
authors.lastname: Iatrou
authors.forename: Luc
authors.initials: L
authors.lastname: Swevers
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017124:Conserved Sequence; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004440:Ecdysone / Q000819:agonists* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D017930:Genes, Reporter / Q000235:genetics*; D006801:Humans; D006834:Hydrazines / Q000737:chemistry / Q000494:pharmacology; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D017434:Protein Structure, Tertiary; D021281:Quantitative Structure-Activity Relationship; D011987:Receptors, Steroid / Q000819:agonists / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: D006834:Hydrazines; D008024:Ligands; D011987:Receptors, Steroid; C055469:ecdysone receptor; C029424:hydrazine; D004440:Ecdysone