Ethylene glycol: Evidence of glucuronidation in vivo shown by analysis of clinical toxicology samples.

In the search for improved laboratory methods for the diagnosis of ethylene glycol poisoning, the in vivo formation of a glucuronide metabolite of ethylene glycol was hypothesized. Chemically pure standards of the ß-O-glucuronide of ethylene glycol (EG-GLUC) and a deuterated analog (d4 -EG-GLUC) were synthesized. A high-performance liquid chromatography and tandem mass spectrometry method for determination of EG-GLUC in serum after ultrafiltration was validated. Inter-assay precision (%RSD) was 3.9% to 15.1% and inter-assay %bias was -2.8% to 12.2%. The measuring range was 2-100 µmol/L (0.48-24 mg/L). Specificity testing showed no endogenous amounts in routine clinical samples (n = 40). The method was used to analyze authentic, clinical serum samples (n = 31) from patients intoxicated with ethylene glycol. EG-GLUC was quantified in 15 of these samples, with a mean concentration of 6.5 µmol/L (1.6 mg/L), ranging from 2.3 to 15.6 µmol/L (0.55 to 3.7 mg/L). In five samples, EG-GLUC was detected below the limit of quantification (2 µmol/L) and it was below the limit of detection in 11 samples (1 µmol/L). Compared to the millimolar concentrations of ethylene glycol present in blood after intoxications and potentially available for conjugation, the concentrations of EG-GLUC found in clinical serum samples are very low, but comparable to concentrations of ethyl glucuronide after medium dose ethanol intake. In theory, EG-GLUC has a potential value as a biomarker for ethylene glycol intake, but the pharmacokinetic properties, in vivo/vitro stability and the biosynthetic pathways of EG-GLUC must be further studied in a larger number of patients and other biological matrices.

Indoloazepinone-Constrained Oligomers as Cell-Penetrating and Blood-Brain-Barrier-Permeating Compounds.

Non-cationic and amphipathic indoloazepinone-constrained (Aia) oligomers have been synthesized as new vectors for intracellular delivery. The conformational preferences of the [l-Aia-Xxx]n oligomers were investigated by circular dichroism (CD) and NMR spectroscopy. Whereas Boc-[l-Aia-Gly]2,4 -OBn oligomers 12 and 13 and Boc-[l-Aia-ß3 -h-l-Ala]2,4 -OBn oligomers 16 and 17 were totally or partially disordered, Boc-[l-Aia-l-Ala]2 -OBn (14) induced a typical turn stabilized by C5 - and C7 -membered H-bond pseudo-cycles and aromatic interactions. Boc-[l-Aia-l-Ala]4 -OBn (15) exhibited a unique structure with remarkable T-shaped π-stacking interactions involving the indole rings of the four l-Aia residues forming a dense hydrophobic cluster. All of the proposed FITC-6-Ahx-[l-Aia-Xxx]4 -NH2 oligomers 19-23, with the exception of FITC-6-Ahx-[l-Aia-Gly]4 -NH2 (18), were internalized by MDA-MB-231 cells with higher efficiency than the positive references penetratin and Arg8 . In parallel, the compounds of this series were successfully explored in an in vitro blood-brain barrier (BBB) permeation assay. Although no passive diffusion permeability was observed for any of the tested Ac-[l-Aia-Xxx]4 -NH2 oligomers in the PAMPA model, Ac-[l-Aia-l-Arg]4 -NH2 (26) showed significant permeation in the in vitro cell-based human model of the BBB, suggesting an active mechanism of cell penetration.

Synthesis and extended activity of triazole-containing macrocyclic protease inhibitors.

Peptide-derived protease inhibitors are an important class of compounds with the potential to treat a wide range of diseases. Herein, we describe the synthesis of a series of triazole-containing macrocyclic protease inhibitors pre-organized into a ß-strand conformation and an evaluation of their activity against a panel of proteases. Acyclic azido-alkyne-based aldehydes are also evaluated for comparison. The macrocyclic peptidomimetics showed considerable activity towards calpain II, cathepsin L and S, and the 20S proteasome chymotrypsin-like activity. Some of the first examples of highly potent macrocyclic inhibitors of cathepsin S were identified. These adopt a well-defined ß-strand geometry as shown by NMR spectroscopy, X-ray analysis, and molecular docking studies.