Prof. Harald Sitte

Investigation of the molecular transport mechanism of monoamine transporters by application of new psychoactive substances.
The investigation of the transport function of monoamine transporters on a molecular model is the prime goal of this project. We will achieve this goal by assessing the activity of different compounds classified as new psychoactive substances (NPS) in either racemic or enantiomer-selective form and possibly also metabolites thereof. Most recently, the first hybrid compounds have been characterized within the framework of this doctoral program (Saha et al., Neuropsychopharmacology, 2014). Hence, the anticipated work in this project proposal will consist of biochemical tracer flux assays (radioactive ligand uptake and release assays), electrophysiology to determine the compound-induced currents and determination of the binding affinities by determination of the kon- and koff-rates. Finally, the student will be expected to work in close collaboration with the two groups of Gerhard Ecker (homology modeling/docking) and Marko Mihovilovic (chemical synthesis). The final goal is to investigate the molecular transport mechanism of monoamine transporters (MAT) by application of new psychoactive substances. These MATs include the transporters for dopamine, norepinephrine and serotonin. Pilot experiments have shown that enantioenriched methcathinones act as inhibitors of transporters. In a 5-HT uptake inhibition assay e.g. (R)- and (S)-mephedrone showed significantly different EC50 values, thus supporting our initial hypothesis.

Workplan: In this project, the methods for the examination of enantioselective and regioselective methcathinones are already established, all radiotracer flux measurements, electrophysiological current detection and imaging techniques (the student will also perform mutagenesis to change selected amino acids and visualize the resulting mutants to detect cell surface expression).
Networking: The project is a joint effort of the Mihovilovic, Sitte and Ecker labs and thus three PhD students will be involved. Compound development and chemical charaterization of compounds will be performed at the Mihovilovic lab. The Ecker group will conduct docking studies of the target compounds at the transporter protein.

Saha K, Partilla JS, Lehner KR, Seddik A, Stockner T, Holy M, Sandtner W, Ecker GF, Sitte HH, Baumann MH. 'Second-Generation' Mephedrone Analogs, 4-MEC and 4-MePPP, Differentially Affect Monoamine Transporter Function. Neuropsychopharmacology. 2014 Dec 15. doi: 10.1038/npp.2014.325. Epub ahead of print
Selected paper by the Faculty of 1000.