Virtual screening for novel openers of pancreatic K(ATP) channels.

Ligand-based virtual screening approaches were applied to search for new chemotype KCOs activating Kir6.2/SUR1 KATP channels. A total of 65 208 commercially available compounds, extracted from the ZINC archive, served as database for screening. In a first step, pharmacokinetic filtering via VolSurf reduced the initial database to 1913 compounds. Afterward, six molecules were selected as templates for similarity searches: similarity scores, obtained toward these templates, were calculated with the GRIND, FLAP, and TOPP approaches, which differently encode structural information into potential pharmacophores. In this way, we obtained 32 hit candidates, 16 via GRIND and eight each via FLAP and TOPP. For biological testing of the hit candidates, their effects on membrane potentials in HEK 293 cells expressing Kir6.2/SUR1 were studied. GRIND, FLAP, and TOPP all yielded hits, but no method top-ranked all the actives. Thus, parallel application of different approaches probably improves hit detection.

New 3-alkylamino-4H-thieno-1,2,4-thiadiazine 1,1-dioxide derivatives activate ATP-sensitive potassium channels of pancreatic beta cells.

Compound 1a (NN414) is a potent opener of Kir6.2/SUR1 K(ATP) channels. Compound 1a inhibits insulin release in vitro and in vivo and preserves beta cell function in preclinical animal models suggesting that such a compound could find use in treatment or prevention of type 1 and type 2 diabetes. The crystal structure and a convergent synthesis of 1a are presented together with a range of new analogues of 1a. Several compounds, e.g., 6-chloro-3-(1-methyl-1-phenylethyl)amino-4H-thieno[3,2-e]-1,2,4-thiadiazine 1,1-dioxide (1h), were found to be potent openers of Kir6.2/SUR1 K(ATP) channels and were able to suppress glucose-stimulated insulin release from rat islets in vitro (EC(50) = 0.04 +/- 0.01 muM) and in vivo after intravenous or peroral administration to hyperinsulinemic obese Zucker rats (ED(50) = 4.0 mg/kg). Structural modifications of this series of K(ATP) channel openers have provided compounds with promising pharmacokinetic properties indicating that brief periods of beta cell rest can be achieved.

Establishment and application of in vitro membrane potential assays in cell lines with endogenous or recombinant expression of ATP-sensitive potassium channels (Kir6.2/SUR1) using a fluorescent probe kit.

The flow of current through the adenosine triphosphate (ATP)-sensitive potassium channel (K(ATP)) of the isoform Kir6.2/SUR1 regulates the resting membrane potential in the pancreatic beta-cell. In combination with the cellular glucose metabolism, it is an important minute-to-minute regulator of insulin secretion and whole-body glucose homeostasis. The same K(ATP) isoform is further reported to be present in glucagon-secreting alpha-cells, intestinal L-cells, and glucose-responsive neurons in the hypothalamus. All in all, this makes Kir6.2/SUR1 an interesting drug target. Using a commercially available fluorescent membrane potential probe kit and a conventional 96-well fluorescence plate reader, the authors have developed and established qualitative membrane potential assays used to screen for potassium channel closers (KCCs) and openers (KCOs) in insulin- and glucagon-secreting cell lines as well as in cells with recombinant expression of the human Kir6.2/SUR1 channel complex. Both glucose- and KCC-induced depolarization could be demonstrated. The magnitudes of these responses and KCO-induced repolarization at high glucose displayed some variation between the different cell lines but a similar rank order of test compounds. Some cell types required the presence of a KCC, such as tolbutamide, to display significant effects of KCOs. The authors find that robust and reliable functional in vitro assays compatible with medium-throughput screening and high-throughput screening can be developed as a base for finding new, more potent, and isoform-selective KCCs and KCOs.