Multi-generational pharmacophore modeling for ligands to the cholane steroid-recognition site in the β1 modulatory subunit of the BKCa channel

Abstract

Large conductance, voltage- and Ca2+-gated K+ (BKCa) channels play a critical role in smooth muscle contractility and thus represent an emerging therapeutic target for drug development to treat vascular disease, gastrointestinal, bladder and uterine disorders. Several compounds are known to target the ubiquitously expressed BKCa channel-forming α subunit. In contrast, just a few are known to target the BKCa modulatory β1 subunit, which is highly expressed in smooth muscle and scarce in most other tissues. Lack of available high-resolution structural data makes structure-based pharmacophore modeling of β1 subunit-dependent BKCa channel activators a major challenge. Following recent discoveries of novel BKCa channel activators that act via β1 subunit recognition, we performed ligand-based pharmacophore modeling that led to the successful creation and fine-tuning of a pharmacophore over several generations. Initial models were developed using physiologically active cholane steroids (bile acids) as template. However, as more compounds that act on BKCa β1 have been discovered, our model has been refined to improve accuracy. Database searching with our best-performing model has uncovered several novel compounds as candidate BKCa β1 subunit ligands. Eight of the identified compounds were experimentally screened and two proved to be activators of recombinant BKCa β1 complexes. One of these activators, sobetirome, differs substantially in structure from any previously reported activator.

Calcium- and voltage-gated BK channels in vascular smooth muscle

“…Therefore, vascular SM BK channel activators are effective vasodilators, as reviewed in [93] while genetic ablation of the BK β1 subunit (e.g., in the KCNMB −/− mouse) is associated with increased myogenic tone [18]. Finally, β1 subunits embolden BK channels with distinct pharmacology, including channel activation by 17β-estradiol [180], micromolar levels of hydrophobic cholanes and their nonsteroidal analogs [22,23,29,132], leukotriene B4 [24], dehydrosoyasaponin-I [131], hydroclorothiazide [127], equol [203], tungstate [57], increased sensitivity to PIP 2 [179] and DHA [74,75], and ethanol-induced reduction of channel activity at low μM Ca 2+ ic [94,95]. The role of SM BK channels as pharmaco-therapeutic targets has been recently reviewed elsewhere [93,219].…”

Celastrol Dilates and Counteracts Ethanol-Induced Constriction of Cerebral Arteries

“…ligands such as bile acids (Bukiya et al, 2011;Dopico et al, 2002) and the synthetic compound termed "HENA" (Bukiya et al, 2013). Indeed, celastrol satisfies major pharmacophore criteria for β 1 -containing BK channels (McMillan et al, 2014). Overall, celastrol maximal vasodilation ranges from 8-12% increase over pre-drug values, when we consider all systems here studied: de-endothelialized and intact isolated MCA segments, and cranial window MCA measurements in live rats.…”

“…Celastrol also exerts neuroprotection, which has been linked to its antioxidant and anti-inflammatory activities (Allison et al, 2001;Cleren et al, 2005;Kiaei et al, 2005;Paris et al, 2010). Remarkably, our group has identified that celastrol fits pharmacophore criteria for activation of BKs that include regulatory subunits abundant in cerebrovascular SM, and has further demonstrated that ex-vivo exposure of these recombinant heteromeric BKs in a heterologous expression system could be activated by celastrol (McMillan et al, 2014). The aim of this study is to determine whether 1) celastrol dilates cerebral arteries both ex-vivo and in-vivo; 2) this action requires the vascular endothelium or, rather, involves SM targets; 3) celastrol-induced cerebral dilation specifically involves BKs; 4) celastrol is able to counteract cerebral artery constriction evoked by ethanol concentrations found in blood during binge drinking.…”

Differential distribution and functional impact of BK channel beta1 subunits across mesenteric, coronary, and different cerebral arteries of the rat

“…4, 5 and 6) and BK β1 mRNA levels (Fig. 7) when compared to those of BA or CA, 2) rat MCA is a widely used experimental model to address human cerebrovascular pathophysiology and pharmacology [3,26,33,88], and 3) we conducted all our previous studies with smooth muscle BK β1-targeting vasodilators (LCA itself, other cholanes, LTB4 and HENA) using rat MCA [15–17,19,63,74]. BK β1 protein overexpression was confirmed by Western blotting (Fig.…”