Bundling arrows: improving translational CNS drug development by integrated PK/PD-metabolomics.

INTRODUCTION: Diseases of the Central Nervous System (CNS) affect millions of people worldwide, with the number of people affected quickly growing. Unfortunately, the successful development of CNS-acting drugs is less than 10%, and this is attributed to the complexity of the CNS, unexpected side effects, difficulties in penetrating the blood-brain barrier and lack of biomarkers. Areas covered: Herein, the authors first review how pharmacokinetic/pharmacodynamic (PK/PD) models are designed to predict the dose-dependent time course of effect, and how they are used to translate drug effects from animal to man. Then, the authors discuss how pharmacometabolomics gives insight into system-wide pharmacological effects and why it is a promising method to study interspecies differences. Finally, the authors advocate the application of PK/PD-metabolomics modeling to advance translational CNS drug development by discussing its opportunities and challenges. Expert opinion: It is envisioned that PK/PD-metabolomics will increase our understanding of CNS drug effects and improve translational CNS drug development, thereby increasing success rates. The successful future development of this concept will require multi-level and longitudinal biomarker evaluation over a large dose range, multi-tissue biomarker evaluation, and the generation of a proof of principle by application to multiple CNS drugs in multiple species.

Multivariate pharmacokinetic/pharmacodynamic (PKPD) analysis with metabolomics shows multiple effects of remoxipride in rats.

The study of central nervous system (CNS) pharmacology is limited by a lack of drug effect biomarkers. Pharmacometabolomics is a promising new tool to identify multiple molecular responses upon drug treatment. However, the pharmacodynamics is typically not evaluated in metabolomics studies, although being important properties of biomarkers. In this study we integrated pharmacometabolomics with pharmacokinetic/pharmacodynamic (PKPD) modeling to identify and quantify the multiple endogenous metabolite dose-response relations for the dopamine D2 antagonist remoxipride. Remoxipride (vehicle, 0.7 or 3.5mg/kg) was administered to rats. Endogenous metabolites were analyzed in plasma using a biogenic amine platform and PKPD models were derived for each single metabolite. These models were clustered on basis of proximity between their PKPD parameter estimates, and PKPD models were subsequently fitted for the individual clusters. Finally, the metabolites were evaluated for being significantly affected by remoxipride. In total 44 metabolites were detected in plasma, many of them showing a dose dependent decrease from baseline. We identified 6 different clusters with different time and dose dependent responses and 18 metabolites were revealed as potential biomarker. The glycine, serine and threonine pathway was associated with remoxipride pharmacology, as well as the brain uptake of the dopamine and serotonin precursors. This is the first time that pharmacometabolomics and PKPD modeling were integrated. The resulting PKPD cluster model described diverse pharmacometabolomics responses and provided a further understanding of remoxipride pharmacodynamics. Future research should focus on the simultaneous pharmacometabolomics analysis in brain and plasma to increase the interpretability of these responses.

Microwave irradiation in label-detection for diagnostic DNA-in situ hybridization.

A DNA-in situ hybridization protocol was adapted for application to sections of routinely processed paraffin embedded material. This protocol was developed previously for detecting DNA-virus infected cells in whole cell preparations and employs biotinylated DNA as probe. Three different biotin detection methods were optimized and applied. The first uses streptavidin and a biotinylated complex of alkaline phosphatase, the second consists of an immunogold-silver staining, and the third of a peroxidase technique using a silver amplification. The alkaline phosphatase method was the most rapid, and as sensitive as the immunogold-silver staining. The peroxidase method was the most sensitive. Microwave irradiation was applied to the different incubation steps of these three detection methods. Short incubations with microwave irradiation gave results comparable to those obtained with conventional incubations, when streptavidin, antibiotin, complexed alkaline phosphatase, or gold labelled goat antirabbit were used. It was thus shown that microwave irradiation creates the possibility of a very rapid label-detection for nonradioactive DNA-in situ hybridization.

Fatty acid acylation of viral proteins in murine hepatitis virus-infected cells. Brief report.

The fatty acid acylation of the cell-associated virus-specific proteins of mouse hepatitis virus (A 59-strain) was studied. 3H-palmitate label was associated with E2, one of the two virion glycoproteins and its intracellular precursor gp 150. A 110 K protein, the unglycosylated apoprotein of gp 150, accumulated by tunicamycin treatment, also incorporated radiolabeled palmitic acid. The addition of fatty acid to the MHV-A 59 E 2 protein is therefore not dependent on glycosylation.