Discovery and characterization of a potent activator of the BKCa channel that relives overactive bladder syndrome in rats

The large-conductance Ca2+-activated K+ channel (BKCa channel) is involved in repolarizing the membrane potential and has a variety of cellular functions. The BKCa channel is highly expressed in bladder smooth muscle and mediates muscle relaxation. Compounds that activate the BKCa channel have therapeutic potential against pathological symptoms associated with the overactivity of bladder smooth muscle. In this regard, we screened a chemical library of 9938 compounds to identify novel BKCa channel activators. A cell-based fluorescence assay identified a structural family of compounds containing a common tricyclic quinazoline ring that activated the BKCa channel. The most potent compound TTQC-1 (7-bromo-N-(3-methylphenyl)-5-oxo-1-thioxo-4,5-dihydro [1,3]thiazolo[3,4-a]quinazoline-3-carboxamide) directly and reversibly activated the macroscopic current of BKCa channels expressed in Xenopus oocytes from both sides of the cellular membrane. TTQC-1 increased the maximum conductance and shifted the half activation voltage to the left. The apparent half-maximal effective concentration and dissociation constant were 2.8 mu M and 7.95 mu M, respectively. TTQC-1 delayed the kinetics of channel deactivation without affecting channel activation. The activation effects were observed over a wide range of intracellular Ca2+ concentrations and dependent on the co-expression of beta 1 and beta 4 auxiliary subunits, which are highly expressed in urinary bladder. In the isolated smooth muscle cells of rat urinary bladder, TTQC-1 increased the K+ currents which can be blocked by iberiotoxin. Finally, oral administration of TTQC-1 to hypertensive rats decreased the urination frequency. Therefore, TTQC-1 is a BKCa channel activator with a novel structure that is a potential therapeutic candidate for BKCa channel-related diseases, such as overactive bladder syndrome.N