Design and Evaluation of [18F]CHDI-650 as a Positron Emission Tomography Ligand to Image Mutant Huntingtin Aggregates.

Therapeutic interventions are being developed for Huntington's disease (HD), a hallmark of which is mutant huntingtin protein (mHTT) aggregates. Following the advancement to human testing of two [11C]-PET ligands for aggregated mHTT, attributes for further optimization were identified. We replaced the pyridazinone ring of CHDI-180 with a pyrimidine ring and minimized off-target binding using brain homogenate derived from Alzheimer's disease patients. The major in vivo metabolic pathway via aldehyde oxidase was blocked with a 2-methyl group on the pyrimidine ring. A strategically placed ring-nitrogen on the benzoxazole core ensured high free fraction in the brain without introducing efflux. Replacing a methoxy pendant with a fluoro-ethoxy group and introducing deuterium atoms suppressed oxidative defluorination and accumulation of [18F]-signal in bones. The resulting PET ligand, CHDI-650, shows a rapid brain uptake and washout profile in non-human primates and is now being advanced to human testing.

Discovery of Novel UDP-N-Acetylglucosamine Acyltransferase (LpxA) Inhibitors with Activity against Pseudomonas aeruginosa.

This study describes a novel series of UDP-N-acetylglucosamine acyltransferase (LpxA) inhibitors that was identified through affinity-mediated selection from a DNA-encoded compound library. The original hit was a selective inhibitor of Pseudomonas aeruginosa LpxA with no activity against Escherichia coli LpxA. The biochemical potency of the series was optimized through an X-ray crystallography-supported medicinal chemistry program, resulting in compounds with nanomolar activity against P. aeruginosa LpxA (best half-maximal inhibitory concentration (IC50) <5 nM) and cellular activity against P. aeruginosa (best minimal inhibitory concentration (MIC) of 4 µg/mL). Lack of activity against E. coli was maintained (IC50 > 20 µM and MIC > 128 µg/mL). The mode of action of analogues was confirmed through genetic analyses. As expected, compounds were active against multidrug-resistant isolates. Further optimization of pharmacokinetics is needed before efficacy studies in mouse infection models can be attempted. To our knowledge, this is the first reported LpxA inhibitor series with selective activity against P. aeruginosa.

Retention-directed and selectivity controlled chromatographic resolution: Rapid post-hoc analysis of DMPK samples to achieve high-throughput LC-MS separation.

High quality chromatographic separation underpins robustness in LC-MS, frequently the analytical method of choice for pharmaceutical drug discovery work. The potential improvements in chromatographic selectivity afforded by serial column coupling (SCC), provide a useful means to enhance the resolution of complex samples. In this work, we present a revised high-throughput form of SCC, in which just two individual mixed phase columns were coupled together and combined with a gradient-optimised, retention-directed ultra-high pressure method to achieve rapid separations, with no further method optimisation necessary. The overall performance was evaluated from an open access DMPK analytical working environment perspective; where in anticipation of bioanalytical or metabolite identification chromatography challenges, or with the knowledge that stronger resolution was required for in-vitro sample analysis, the methodology could be immediately implemented by the analyst. Retention-directed selection of a shallow SCC gradient method was successful in separating peaks throughout the chromatographic window, resulting in a runtime still congruent to high-throughput analyses (3.5 min). In-vitro assay sample interferences were resolved 44-72% of the time, and the overall resolving power for isomeric separations significantly improved against single column comparisons (1.7-fold mean RS improvement). Over a sustained period of time in our laboratory, SCC methods have been used for metabolite identification and bioanalytical samples, where both convenience and effectiveness in solving analytical challenges has been consistently demonstrated. Examples that highlight SCC chromatography, and a guided discussion of the main high-throughput considerations, are included. The technique offers wide applicability, and we would recommend it as a toolbox consideration to the laboratory analyst.

Imaging Mutant Huntingtin Aggregates: Development of a Potential PET Ligand.

Mutant huntingtin (mHTT) protein carrying the elongated N-terminal polyglutamine (polyQ) tract misfolds and forms protein aggregates characteristic of Huntington's disease (HD) pathology. A high-affinity ligand specific for mHTT aggregates could serve as a positron emission tomography (PET) imaging biomarker for HD therapeutic development and disease progression. To identify such compounds with binding affinity for polyQ aggregates, we embarked on systematic structural activity studies; lead optimization of aggregate-binding affinity, unbound fractions in brain, permeability, and low efflux culminated in the discovery of compound 1, which exhibited target engagement in autoradiography (ARG) studies in brain slices from HD mouse models and postmortem human HD samples. PET imaging studies with 11C-labeled 1 in both HD mice and WT nonhuman primates (NHPs) demonstrated that the right-hand-side labeled ligand [11C]-1R (CHDI-180R) is a suitable PET tracer for imaging of mHTT aggregates. [11C]-1R is now being advanced to human trials as a first-in-class HD PET radiotracer.

Lumiracoxib metabolism in male C57bl/6J mice: characterisation of novel in vivo metabolites.

1. The pharmacokinetics and metabolism of lumiracoxib in male C57bl/6J mice were investigated following a single oral dose of 10 mg/kg. 2. Lumiracoxib achieved peak observed concentrations in the blood of 1.26 + 0.51 µg/mL 0.5 h (0.5-1.0) post-dose with an AUCinf of 3.48 + 1.09 µg h/mL. Concentrations of lumiracoxib then declined with a terminal half-life of 1.54 + 0.31 h. 3. Metabolic profiling showed only the presence of unchanged lumiracoxib in blood by 24 h, while urine, bile and faecal extracts contained, in addition to the unchanged parent drug, large amounts of hydroxylated and conjugated metabolites. 4. No evidence was obtained in the mouse for the production of the downstream products of glutathione conjugation such as mercapturates, suggesting that the metabolism of the drug via quinone-imine generating pathways is not a major route of biotransformation in this species. Acyl glucuronidation appeared absent or a very minor route. 5. While there was significant overlap with reported human metabolites, a number of unique mouse metabolites were detected, particularly taurine conjugates of lumiracoxib and its oxidative metabolites.

Reversible C-C bond formation: solid state structure of the aldol-like addition product of adamantanone to a 1,5-diazapentadienyllithium, and its solution state retro-aldol dissociation.

Analysis of crystals of the lithium complex of the tripodal ligand formed upon addition of adamantanone to a 1,5 diazapentadienyllithium complex reveals a long C-C bond which ruptures upon dissolution in non-co-ordinating solvents.