ABSTRACT
Immune-mediated drug-induced liver injury (IMDILI) can be devastating, irreversible, and fatal in the absence of successful transplantation surgery. We present a novel approach that combines the methods of pharmacoepidemiology with in silico molecular modeling to identify specific features in toxic ligands that are associated with clinical features of IMDILI. Specifically, from pharmacovigilance data multivariate logistic regression identified 18 drugs associated with IMDILI (P < 0.00015). Eleven of these drugs, along with their known and proposed metabolites, constituted a training set used to develop a four-point pharmacophore model (sensitivity 75%; specificity 85%). Subsequently, this information was combined with information from immune-pathway reviews and genetic-association studies and complemented with ligand-protein docking simulations to support a hypothesis implicating two putative targets within separate, possibly interacting, immune-system pathways: the major histocompatibility complex within the adaptive immune system and Toll-like receptors (TLRs), in particular TLR-7, which represent pattern recognition receptors of the innate immune system.
ABSTRACT
A direct assay method for use in studies of cyclosporin binding must be highly sensitive and selective since it must be capable of measuring the concentrations encountered in the protein-free matrix. The failure of current HPLC methods to achieve the sensitivity required for binding studies may be attributed to the use of UV detection, which relies on the relatively weak end-absorption of cyclosporin A. A method involving fluorescence derivatization was sought with the aim of increasing HPLC assay sensitivity. A method is described for producing a fluorescent derivative of cyclosporin A, a compound which has no functional groups which are easily derivatized. However, intramolecular rearrangement of cyclosporin A to form its structural isomer, isocyclosporin A, exposes a secondary amine which can be reacted with dansyl chloride to produce a fluorescent derivative. This two-step derivatization procedure was used as the basis of an HPLC fluorescence assay. Although this assay is not sufficiently sensitive to measure concentrations encountered in the protein-free matrix during plasma binding studies, the method does point to the possible development of a more sensitive assay using a derivatizing reagent other than dansyl chloride.
