Inhibition of PDE5A1 guanosine cyclic monophosphate (cGMP) hydrolysing activity by sildenafil analogues that inhibit cellular cGMP efflux.

OBJECTIVES: To determine the ability of 11 sildenafil analogues to discriminate between cyclic nucleotide phosphodiesterases (cnPDEs) and to characterise their inhibitory potencies (Ki values) of PDE5A1-dependent guanosine cyclic monophosphate (cGMP) hydrolysis. METHODS: Sildenafil analogues were identified by virtual ligand screening (VLS) and screened for their ability to inhibit adenosine cyclic monophosphate (cAMP) hydrolysis by PDE1A1, PDE1B1, PDE2A1, PDE3A, PDE10A1 and PDE10A2, and cGMP hydrolysis by PDE5A, PDE6C, PDE9A2 for a low (1 nm) and high concentration (10 µm). Complete IC50 plots for all analogues were performed for PDE5A-dependent cGMP hydrolysis. Docking studies and scoring were made using the ICM molecular modelling software. KEY FINDINGS: The analogues in a low concentration showed no or low inhibition of PDE1A1, PDE1B1, PDE2A1, PDE3A, PDE10A1 and PDE10A2. In contrast, PDE5A and PDE6C were markedly inhibited to a similar extent by the analogues in a low concentration, whereas PDE9A2 was much less inhibited. The analogues showed a relative narrow range of Ki values for PDE5A inhibition (1.2-14 nm). The sildenafil molecule was docked in the structure of PDE5A1 co-crystallised with sildenafil. All the analogues had similar binding poses as sildenafil. CONCLUSIONS: Sildenafil analogues that inhibit cellular cGMP efflux are potent inhibitors of PDE5A and PDE6C.

Novel cGMP efflux inhibitors identified by virtual ligand screening (VLS) and confirmed by experimental studies.

Elevated intracellular levels of cyclic guanosine monophosphate (cGMP) may induce apoptosis, and at least some cancer cells seem to escape this effect by increased efflux of cGMP, as clinical studies have shown that extracellular cGMP levels are elevated in various types of cancer. The human ATP binding cassette (ABC) transporter ABCC5 transports cGMP out of cells, and inhibition of ABCC5 may have cytotoxic effects. Sildenafil inhibits cGMP efflux by binding to ABCC5, and in order to search for potential novel ABCC5 inhibitors, we have identified sildenafil derivates using structural and computational guidance and tested them for the cGMP efflux effect. Eleven compounds from virtual ligand screening (VLS) were tested in vitro, using inside-out vesicles (IOV), for inhibition of cGMP efflux. Seven of 11 compounds predicted by VLS to bind to ABCC5 were more potent than sildenafil, and the two most potent showed K(i) of 50-100 nM.