Abstract

Small-conductance (K_Ca2) and intermediate-conductance (K_Ca3.1) calcium-activated K⁺ channels are voltage-independent and share a common calcium/calmodulin-mediated gating mechanism. Existing positive gating modulators like EBIO, NS309, or SKA-31 activate both K_Ca2 and K_Ca3.1 channels with similar potency or, as in the case of CyPPA and NS13001, selectively activate K_Ca2.2 and K_Ca2.3 channels. We performed a structure-activity relationship (SAR) study with the aim of optimizing the benzothiazole pharmacophore of SKA-31 toward K_Ca3.1 selectivity. We identified SKA-111 (5-methylnaphtho[1,2-d]thiazol-2-amine), which displays 123-fold selectivity for K_Ca3.1 (EC₅₀ 111 ± 27 nM) over K_Ca2.3 (EC₅₀ 13.7 ± 6.9 μM), and SKA-121 (5-methylnaphtho[2,1-d]oxazol-2-amine), which displays 41-fold selectivity for K_Ca3.1 (EC₅₀ 109 nM ± 14 nM) over K_Ca2.3 (EC₅₀ 4.4 ± 1.6 μM). Both compounds are 200- to 400-fold selective over representative K_V (K_V1.3, K_V2.1, K_V3.1, and K_V11.1), Na_V (Na_V1.2, Na_V1.4, Na_V1.5, and Na_V1.7), as well as Ca_V1.2 channels. SKA-121 is a typical positive-gating modulator and shifts the calcium-concentration response curve of K_Ca3.1 to the left. In blood pressure telemetry experiments, SKA-121 (100 mg/kg i.p.) significantly lowered mean arterial blood pressure in normotensive and hypertensive wild-type but not in KCa3.1^−/− mice. SKA-111, which was found in pharmacokinetic experiments to have a much longer half-life and to be much more brain penetrant than SKA-121, not only lowered blood pressure but also drastically reduced heart rate, presumably through cardiac and neuronal K_Ca2 activation when dosed at 100 mg/kg. In conclusion, with SKA-121, we generated a K_Ca3.1-specific positive gating modulator suitable for further exploring the therapeutical potential of K_Ca3.1 activation.