Christopher G. Boissard

Targeting acute ischemic stroke with a calcium-sensitive opener of maxi-K potassium channels

During ischemic stroke, neurons at risk are exposed to pathologically high levels of intracellular calcium (Ca++), initiating a fatal biochemical cascade. To protect these neurons, we have developed openers of large-conductance, Ca++-activated (maxi-K or BK) potassium channels, thereby augmenting an endogenous mechanism for regulating Ca++ entry and membrane potential. The novel fluoro-oxindoles BMS-204352 and racemic compound 1 are potent, effective and uniquely Ca++-sensitive openers of maxi-K channels. In rat models of permanent large-vessel stroke, BMS-204352 provided significant levels of cortical neuroprotection when administered two hours after the onset of occlusion, but had no effects on blood pressure or cerebral blood flow. This novel approach may restrict Ca++ entry in neurons at risk while having minimal side effects.

Formation of intermediate-conductance calcium-activated potassium channels by interaction of Slack and Slo subunits

Large-conductance calcium-activated potassium channels (maxi-K channels) have an essential role in the control of excitability and secretion. Only one gene Slo is known to encode maxi-K channels, which are sensitive to both membrane potential and intracellular calcium. We have isolated a potassium channel gene called Slack that is abundantly expressed in the nervous system. Slack channels rectify outwardly with a unitary conductance of about 25–65 pS and are inhibited by intracellular calcium. However, when Slack is co-expressed with Slo, channels with pharmacological properties and single-channel conductances that do not match either Slack or Slo are formed. The Slack/Slo channels have intermediate conductances of about 60–180 pS and are activated by cytoplasmic calcium. Our findings indicate that some intermediate-conductance channels in the nervous system may result from an interaction between Slack and Slo channel subunits.

The synthesis and characterization of BMS-204352 (MaxiPost) and related 3-fluorooxindoles as openers of maxi-K potassium channels

3-Aryl-3-fluorooxindoles can be efficiently synthesized in two steps by the addition of an aryl Grignard to an isatin, followed by treatment with DAST. Oxindole 1 (BMS-204352; MaxiPost) can be isolated using chiral HPLC or prepared by employing chiral resolution. Cloned maxi-K channels are opened by 1, which demonstrates a brain/plasma ratio >9 in rats.

The synthesis and structure-activity relationships of 4-aryl-3-aminoquinolin-2-ones: A new class of calcium-dependent, large conductance, potassium (maxi-K) channel openers targeted for post-stroke neuroprotection

A series of 4-aryl-3-aminoquinoline-2-one derivatives was synthesized and evaluated as activators of the cloned maxi-K channel mSlo (hSlo) expressed in Xenopus laevis oocytes using electrophysiological methods. A brain penetrable activator of maxi-K channels was identified and shown to be significantly active in the MCAO model of stroke.

Discovery of a novel class of BK channel openers : Enantiospecific synthesis and BK channel opening activity of 3-(5-chloro-2-hydroxyphenyl)-1,3-dihydro-3-hydroxy-6-(trifluoromethyl)-2H-indol-2-one

Abstract 3-Aryl-3-hydroxyindol-2-ones have been identified as a novel class of BK channel openers. Synthesis of both racemic and chiral 3-aryl-3-hydroxyindolones is described along with their electrophysiological evaluation as activators of the cloned BK channel mSlo expressed in Xenopus laevis oocytes. The preliminary SAR data indicate the importance of both an electron-withdrawing substituent on the oxindole nucleus and the phenolic hydroxyl for expression of BK channel opening properties. Moreover, the dependence of BK channel opening activity on the absolute configuration of the chiral center in this pharmacophore has been demonstrated.

Pharmacological activation and inhibition of Slack (Slo2.2) channels

The Slack (Sequence like a calcium-activated K channel) (Slo2.2) gene is abundantly expressed in the mammalian brain and encodes a sodium-activated K+ (KNa) channel. Although the specific roles of Slack channel subunits in neurons remain to be identified, they may play a role in the adaptation of firing rate and in protection against ischemic injury. In the present study, we have generated a stable cell line expressing the Slack channel, and have analyzed the pharmacological properties of these channels in these cells and in Xenopus oocytes. Two known blockers of KNa channels, bepridil and quinidine, inhibited Slack currents in a concentration-dependent manner and decreased channel activity in excised membrane patches. The inhibition by bepridil was potent, with an IC50 of 1.0 microM for inhibition of Slack currents in HEK cells. In contrast, bithionol was found to be a robust activator of Slack currents. When applied to the extracellular face of excised patches, bithionol rapidly induced a reversible increase in channel opening, suggesting that it acts on Slack channels relatively directly. These data establish an important early characterization of agents that modulate Slack channels, a process essential for the experimental manipulation of Slack currents in neurons.

The synthesis and structure-activity relationships of 1,3-diaryl 1,2,4-(4H)-triazol-5-ones: a new class of calcium-dependent, large conductance, potassium (maxi-K) channel opener targeted for urge urinary incontinence.

A series of 1,3-diaryl 1,2,4-(4H)-triazol-5-ones was prepared and shown by electrophysiological analysis to activate a cloned maxi-K channel mSlo (or hSlo) expressed in Xenopus laevis oocytes. The effects of these structurally novel maxi-K channel openers on bladder contractile function were studied in vitro using isolated rat bladder strips pre-contracted with carbachol. Several 1,3-diaryl 1,2,4-(4H)-triazol-5-one derivatives were found to be potent smooth muscle relaxants but this activity did not completely correlate with maxi-K channel opening.

Synthesis and excitatory amino acid pharmacology of some novel quinoxalinediones

Abstract The synthesis and amino acid pharmacology of twelve N-substituted quinoxalinediones is reported. In particular, compounds 4a and 4b show significant antagonism at both the AMPA and glycine-site NMDA receptors. The functional antagonism of 4a has been demonstrated.

The discovery of novel openers of Ca2+-dependent large-conductance potassium channels: Pharmacophore search and physiological evaluation of flavonoids

Abstract Three-dimensional database searching based on the pharmacophore model of the known maxi-K opener NS-004 ( 1 ) and electrophysiological evaluation led to the discovery of flavonoids ( 3c and 3d ) as openers of the cloned maxi-K channel m Slo expressed in Xenopus laevis oocytes. The cyclic constrained flavonoids were found to be more efficacious than the acyclic phloretin.

N-Benzylated benzimidazol-2-one derivatives: activators of large-conductance Ca2+-dependent K+ channels

Abstract A series of benzimidazol-2-ones structurally homologous to the known maxi-K opener NS-004 (2) were synthesized and evaluated by electrophysiological techniques as openers of the cloned maxi-K channel m Slo expressed in Xenopus laevis oocytes. The structure-activity relationships reveal tolerance in the topological relationship between the heterocycle and the phenol hydroxy and indicate the importance of an electron withdrawing substituent on the heterocycle for expression of maxi-K opening properties. The most efficacious activators of this channel were the 5-Cl derivative 4f and the 5-NO 2 analogue 4i .