Nicholas C. O. Tomkinson

The Pregnane X Receptor: A Promiscuous Xenobiotic Receptor That Has Diverged during Evolution

Transcription of genes encoding cytochrome P450 3A (CYP3A) monooxygenases is induced by a variety of xenobiotics and natural steroids. There are marked differences in the compounds that induce CYP3A gene expression between species. Recently, the mouse and human pregnane X receptor (PXR) were shown to be activated by compounds that induce CYP3A expression. However, most studies of CYP3A regulation have been performed using rabbit and rat hepatocytes. Here, we report the cloning and characterization of PXR from these two species. PXR is remarkably divergent between species, with the rabbit, rat, and human receptors sharing only approximately 80% amino acid identity in their ligand-binding domains. This sequence divergence is reflected by marked pharmacological differences in PXR activation profiles. For example, the macrolide antibiotic rifampicin, the antidiabetic drug troglitazone, and the hypocholesterolemic drug SR12813 are efficacious activators of the human and rabbit PXR but have little activity on the rat and mouse PXR. Conversely, pregnane 16alpha-carbonitrile is a more potent activator of the rat and mouse PXR than the human and rabbit receptor. The activities of xenobiotics in PXR activation assays correlate well with their ability to induce CYP3A expression in primary hepatocytes. Through the use of a novel scintillation proximity binding assay, we demonstrate that many of the compounds that induce CYP3A expression bind directly to human PXR. These data establish PXR as a promiscuous xenobiotic receptor that has diverged during evolution.

Identification of peroxisome proliferator-activated receptor ligands from a biased chemical library

BACKGROUND The peroxisome proliferator-activated receptors (PPARs) were cloned as orphan members of the nuclear receptor superfamily of transcription factors. The identification of subtype-selective ligands for PPARalpha and PPARgamma has led to the discovery of their roles in the regulation of lipid metabolism and glucose homeostasis. No subtype-selective PPARdelta ligands are available and the function of this subtype is currently unknown. RESULTS A three-component library was designed in which one of the monomers was biased towards the PPARs and the other two monomers were chosen to add chemical diversity. Synthesis and screening of the library resulted in the identification of pools with activity on each of the PPAR subtypes. Deconvolution of the pools with the highest activity on PPARdelta led to the identification of GW 2433 as the first high-affinity PPARdelta ligand. [3H]GW 2433 is an effective radioligand for use in PPARdelta competition-binding assays. CONCLUSIONS The synthesis of biased chemical libraries is an efficient approach to the identification of lead molecules for members of sequence-related receptor families. This approach is well suited to the discovery of small-molecule ligands for orphan receptors.

A public-private partnership to unlock the untargeted kinome

Chemical probes are urgently needed to functionally annotate hitherto-untargeted kinases and stimulate new drug discovery efforts to address unmet medical needs. The size of the human kinome combined with the high cost associated with probe generation severely limits access to new probes. We propose a large-scale public-private partnership as a new approach that offers economies of scale, minimized redundancy and sharing of risk and cost.

Secondary and Primary Amine Catalysts for Iminium Catalysis

Formation of iminium ions from the condensation of chiral secondary or primary amines with alpha,beta-unsaturated aldehydes or ketones can be used as an effective platform for the acceleration of a wide variety of catalytic asymmetric cycloaddition and conjugate addition reactions. The reversible formation of the active iminium ion species simulates the pi-electronics and equilibrium dynamics traditionally associated with Lewis acid activation of alpha,beta-unsaturated carbonyl compounds lowering the energy level of the LUMO associated with the pi-system and activating subsequent reaction. Importantly, these iminium ion catalysed processes offer the opportunity to conduct reactions in the presence of both moisture and air greatly adding to the practicality and general applicability of the chemistry described. Proposed catalytic cycles and transition state models for the induction of asymmetry provide reliable and robust predictive tools for the outcome of reactions and high functional group tolerance suggests this class of transformation will have broad application in the arena of synthetic organic chemistry as the area matures. This review describes the rapid expansion of iminium ion catalysis over recent years from its conceptual introduction to the development of a whole new arsenal of highly practical and effective methods with which to approach challenging and fundamental bond construction processes.

Solution phase, solid state, and theoretical investigations on the MacMillan imidazolidinone.

A combination of solution phase NMR, X-ray crystallographic studies, and DFT calculations provide a consistent structural conformation for iminium ions derived from the MacMillan imidazolidinone.

Differential activity of rosiglitazone enantiomers at PPARγ

Anal. of the enantiomers of rosiglitazone in a PPARγ binding assay suggests that the (S)-(-)- isomer is responsible for the antidiabetic activity.

Kinetics of iminium ion catalysis.

To a stirred solution of cinnamaldehyde (118 mg, 1.42 mmol, 122 ?l) in methanol (2ml), trifluoromethylpyrrolidine (198 mg, 1.42 mmol) was added and allowed to stir at ambient temperature for 2 minutes. After this time, hexafluorophosphoric solution in H2O (60% w/w) (345 mg, 1.42 mmol, 210 ?l) was added which immediately formed a yellow precipitate on addition. The excess solvent was removed in vacuo and the crude mixture was recrystallised from hot methanol to give 6 (493 mg, 87%) as an off white solid; mp 199-201 oC; ?max(Nujol) 2923, 2853, 1621, 1591, 1456, 1376 cm-1; 1H NMR (400MHz, CD3CN) ? 8.64 (d, J = 10.6 Hz, 1H), 8.12 (d, J = 15.2 Hz, 1H), 7.91-7.89 (m, 2H), 7.69-7.67 (m, 1H), 7.60-7.57 (m, 2H), 7.32 (dd, J = 10.6Hz, 15.2Hz, 1H), 5.0-4.96 (m, 1H), 4.24-4.14 (m, 2H), 2.52-2.45 (m, 1H), 2.42-2.27 (m, 3H); 13C NMR (125 MHz, CDCl3) ? 170.8, 165.3, 134.9, 131.2, 129.7, 125.1, 118.4, 66.8 (q, J1CF = 31.3 Hz), 53.3, 24.9, 22.6, 21.7; m/z (APCI) 254 [M–PF6]?+; HRMS (ES) found 254.1152 [M–PF6].+; calculated for C14H15NF3 254.1151.

Copper-catalyzed coupling of hydroxylamines with aryl iodides.

An efficient method for the copper-catalyzed N-arylation of hydroxylamines with aryl iodides is described. A variety of N- and O-functionalized hydroxylamines were transformed in good to excellent yield with a broad range of aryl coupling partners. Methods for the selective deprotection of either the N- or O-substituents for further functionalization are also described.

Metal-free dihydroxylation of alkenes using cyclobutane malonoyl peroxide

Cyclobutane malonoyl peroxide (7), prepared in a single step from the commercially available diacid 6, is an effective reagent for the dihydroxylation of alkenes. Reaction of a chloroform solution of 7 with an alkene in the presence of 1 equiv of water at 40 °C followed by alkaline hydrolysis leads to the corresponding diol (30-84%). With 1,2-disubstituted alkenes, the reaction proceeds with syn-selectivity (3:1 → 50:1). A mechanism consistent with experimental findings is proposed, which is supported by deuterium and oxygen labeling studies and explains the stereoselectivity observed. Alternative reaction pathways that are dependent on the structure of the starting alkene are also described leading to the synthesis of allylic alcohols and γ-lactones.

Iminium ion catalysis: Use of the α-effect in the acceleration of the Diels–Alder reactionElectronic supplementary information (ESI) available: 1H NMR, 13C NMR, IR and MS spectra. See http://www.rsc.org/suppdata/cc/b2/b212239a/

The α-effect can be used in the acceleration of the Diels–Alder reaction between a series of dienes and electron deficient dienophiles using iminium ion catalysis, providing a novel molecular scaffold capable of performing this class of catalytic process.