Identification and characterization of novel small molecule modulators of adenylyl cyclase

Abstract

Adenylyl cyclase (AC) isoforms display distinct regulatory and expression patterns, and have specific roles in fundamental physiological processes. These properties have allowed ACs to be considered as therapeutic targets; however, there are few potent and isoform‐selective small molecule modulators of ACs. The objective of these studies was to identify new AC modulators by developing a high throughput screening assay for small molecule inhibitors of AC2. The NIH Clinical Collections (727 compounds) were screened for inhibition of PMA‐stimulated cAMP accumulation in HEK‐AC2 cells. The most active compounds were prioritized by potency for inhibition of AC2 activity, retention of inhibition in an orthogonal cAMP detection format, and in vitro activity against AC isoforms. Actives were also assessed by examining their effect on PMA‐stimulated ERK1/2 phosphorylation to control for inhibition of AC2 via a direct effect on PKC. Compounds were then tested for AC isoform‐selectivity using additional intact cell assays. Three compounds were identified and validated as AC2 inhibitors with unique selectivity patterns against AC1 and AC5. Subsequent studies used cultured bronchial smooth muscle cells to reveal that the most active compound significantly inhibited AC‐regulated interleukin 6 production, a response that is specifically mediated by AC2. Supported by NIH: MH060397.