Niall M. Hamilton

The in vitro and in vivo enantioselectivity of etomidate implicates the GABAA receptor in general anaesthesia

General anaesthetics exhibiting enantioselectivity afford valuable tools to assess the fundamental mechanisms underlying anaesthesia. Here, we characterised the actions of the R-(+)- and S-(-)-enantiomers of etomidate. In mice and tadpoles, R-(+)-etomidate was more potent (approximately 10-fold) than S-(-)-etomidate in producing loss of the righting reflex. In electrophysiological and radioligand binding assays, the enantiomers of etomidate positively regulated GABAA receptor function at anaesthetic concentrations and with an enantioselectivity paralleling their in vivo activity. GABA-evoked currents mediated by human recombinant GABAA receptors were potentiated by either R-(+)- or S-(-)-etomidate in a manner dependent upon receptor subunit composition. A direct, GABA-mimetic, effect was similarly subunit dependent. Modulation of GABA receptor activity was selective; R-(+)-etomidate inhibited nicotinic acetylcholine, or 5-hydroxytryptamine3 receptor subtypes only at supra-clinical concentrations and ionotropic glutamate receptor isoforms were essentially unaffected. Acting upon reticulothalamic neurones in rat brain slices, R-(+)-etomidate prolonged the duration of miniature IPSCs and modestly enhanced their peak amplitude. S-(-)-etomidate exerted qualitatively similar, but weaker, actions. In a model of locomotor activity, fictive swimming in Xenopus laevis tadpoles, R-(+)- but not S-(-)-etomidate exerted a depressant influence via enhancement of GABAergic neurotransmission. Collectively, these observations strongly implicate the GABAA receptor as a molecular target relevant to the anaesthetic action of etomidate.

Toxoflavins and Deazaflavins as the First Reported Selective Small Molecule Inhibitors of Tyrosyl-DNA Phosphodiesterase II

The recently discovered enzyme tyrosyl-DNA phosphodiesterase 2 (TDP2) has been implicated in the topoisomerase-mediated repair of DNA damage. In the clinical setting, it has been hypothesized that TDP2 may mediate drug resistance to topoisomerase II (topo II) inhibition by etoposide. Therefore, selective pharmacological inhibition of TDP2 is proposed as a novel approach to overcome intrinsic or acquired resistance to topo II-targeted drug therapy. Following a high-throughput screening (HTS) campaign, toxoflavins and deazaflavins were identified as the first reported sub-micromolar and selective inhibitors of this enzyme. Toxoflavin derivatives appeared to exhibit a clear structure-activity relationship (SAR) for TDP2 enzymatic inhibition. However, we observed a key redox liability of this series, and this, alongside early in vitro drug metabolism and pharmacokinetics (DMPK) issues, precluded further exploration. The deazaflavins were developed from a singleton HTS hit. This series showed distinct SAR and did not display redox activity; however low cell permeability proved to be a challenge.

Novel Steroid Inhibitors of Glucose 6-Phosphate Dehydrogenase

Novel derivatives of the steroid DHEA 1, a known uncompetitive inhibitor of G6PD, were designed, synthesized, and tested for their ability to inhibit this dehydrogenase enzyme. Several compounds with approximately 10-fold improved potency in an enzyme assay were identified, and this improved activity translated to efficacy in a cellular assay. The SAR for steroid inhibition of G6PD has been substantially developed; the 3β-alcohol can be replaced with 3β-H-bond donors such as sulfamide, sulfonamide, urea, and carbamate. Improved potency was achieved by replacing the androstane nucleus with a pregnane nucleus, provided a ketone at C-20 is present. For pregnan-20-ones incorporation of a 21-hydroxyl group is often beneficial. The novel compounds generally have good physicochemical properties and satisfactory in vitro DMPK parameters. These derivatives may be useful for examining the role of G6PD inhibition in cells and will assist the future design of more potent steroid inhibitors with potential therapeutic utility.

The Anaesthetic Action and Modulation of GABAA Receptor Activity by the Novel Water-soluble Aminosteroid Org 20599

The anaesthetic profile of a novel water-soluble aminosteroid, Org 20599 [(2 beta, 3 alpha, 5 alpha)-21-chloro-3-hydroxy-2-(4-morpholinyl)pregnan-20-one methanesulphonate], and the ability of the compound to allosterically regulate the activity of the GABAA receptor, have been studied in comparison to the properties of established intravenous general-anaesthetic agents. Intravenously administered Org 20599 produced a rapid onset, short duration loss of the righting reflex in mice. The anaesthetic potency of Org 20599 was comparable to that of the steroids 5 alpha-pregnan-3 alpha-ol-20-one or alphaxalone, and exceeded that of propofol, thiopentone or pentobarbitone. Org 20599 and the reference anaesthetic agents allosterically displaced the binding of [35S]-t-butylbicyclophosphorothionate (TBPS) from GABAA receptors of rat-brain membranes with the order of potency: 5 alpha-pregnan-3 alpha-ol-20-one > Org 20599 > alphaxalone > propofol > thiopentone > pentobarbitone. At human recombinant alpha 1, beta 2, gamma 2L subunit-containing GABAA receptors expressed in Xenopus laevis oocytes, the anaesthetic agents produced a concentration-dependent and reversible potentiation of the peak amplitude of GABA-evoked currents. A similar positive allosteric action of Org 20599 was observed for the GABAA receptors expressed by bovine adrenal chromaffin cells maintained in culture. The rank order of potency in the aforementioned assays was identical to that determined from the displacement of TBPS binding. At concentrations greater than those required for potentiation of GABA, the anaesthetics exhibited GABA-mimetic effects with a rank order of potency that paralleled their modulatory activity. Such direct agonism varied greatly in maximal effect between compounds. The modulatory and direct agonist actions of Org 20599 were additionally confirmed utilizing rat hippocampal neurones in culture. The results indicate Org 20599 to be a potent and short-acting intravenous anaesthetic agent in mice and suggest positive allosteric regulation of GABAA receptor function to be a plausible molecular mechanism of action for the drug.

Structural basis for agonism and antagonism for a set of chemically related progesterone receptor modulators

The progesterone receptor is able to bind to a large number and variety of ligands that elicit a broad range of transcriptional responses ranging from full agonism to full antagonism and numerous mixed profiles inbetween. We describe here two new progesterone receptor ligand binding domain x-ray structures bound to compounds from a structurally related but functionally divergent series, which show different binding modes corresponding to their agonistic or antagonistic nature. In addition, we present a third progesterone receptor ligand binding domain dimer bound to an agonist in monomer A and an antagonist in monomer B, which display binding modes in agreement with the earlier observation that agonists and antagonists from this series adopt different binding modes.

Water-soluble propofol analogues with intravenous anaesthetic activity

Propofol (2,6-diisopropylphenol) is a widely used intravenous anaesthetic that is formulated as an emulsion since it lacks water solubility. We report a range of water-soluble analogues of propofol, containing a para-alkylamino substituent, which retain good intravenous anaesthetic activity in rodents.

A facile synthesis of N-benzylallylglycine

Abstract N -Benzylallylglycine can be prepared in good yield through a one-pot reaction of N -benzylhomoallylamine with glyoxylic acid monohydrate in methanol. The reaction method described requires only mild conditions and avoids the need for purification of the amino acid product.

Novel water soluble 2,6-dimethoxyphenyl ester derivatives with intravenous anaesthetic activity.

A number of water soluble bis-amino-2,6-dimethoxyphenyl ester derivatives were found to exhibit improved anaesthetic activity in mice relative to propofol 1. Of the analogues disclosed, 44 was further profiled in rodents and found to be a superior agent to propofol for the induction and maintenance of anaesthesia.

α-Subunit selective modulators of GABAA receptor function as CNS therapeutics

γ-Aminobutyric acid (GABA) is the major inhibitory neurotransmitter within the CNS. Many drugs, including the benzodiazepines, exert their effects by modulating the GABAA receptor complex, but side effects are common and reflect, in part, poor subunit selectivity. The subunits are arranged into heteropentamers and this produces a rich diversity of GABAA receptor subtypes, which are potential targets for treating a variety of CNS disorders including epilepsy, anxiety and insomnia, as well as for ameliorating deficiencies arising from neurodegeneration, such as cognitive impairment. The identification of new ligands with improved subunit selectivity should reduce or abolish some of the side effects observed with current drugs, such as tolerance, dependence and withdrawal. This article focuses on new ligands that are reported to selectively recognise particular α-subunits of GABAA receptors and may thereby offer improved treatments for CNS disorders. Only patents and literature since 1998 are necessarily included, although some earlier reports and reviews are also cited. Several publications and patent applications since 1998 disclose new compounds that modulate GABAA receptor function without mentioning whether they have α-subunit selectivity; these compounds, like those known to interact with sites on other GABAA subunits (particularly β), are not within the scope of this review.

Therapeutic potential of steroids for CNS disorders

The importance of steroids for modulation of the CNS is now better understood but it is still in its infancy compared with the knowledge that has been accumulated on their endocrine effects. The structural diversity of steroids that exert CNS activity is broad and they modulate their effects, not only via classical steroid receptors (oestrogen, androgen, progestin, glucocorticoid and mineralocorticoid) but also via ion channels, particularly GABAA receptors and glutamate (NMDA) receptors. This review illustrates the therapeutic potential of steroids for treatment of CNS disorders by covering publications and patent literature since 1995.