Description:
<jats:p>Methcathinone (MCAT) is a compound belonging to the class of cathinones which targets monoamine transporters including DAT and SERT. Despite the importance of DAT and SERT as drug targets in several neurological disorders, the key factors underlying the selectivity profiles of their inhibitors are still poorly understood. Recent findings from rat synaptosomes suggested that increasing the volume of the para‐substituent of MCAT results in a swap of the selectivity between human DAT and SERT (1). Docking studies hint towards Ser149 in DAT and Ala169 in SERT as key residues involved in the difference of activity between DAT and SERT (2). The aims of the present biochemical and pharmacological study are to understand (i) which chemical properties (e.g. volume, polarity, or lipophilicity) of the para‐substituent influence the selectivity profile of MCAT between DAT and SERT, and (ii) whether Ser149 in DAT and Ala169 in SERT can be experimentally verified as key residues. Hence, we combined <jats:italic>in silico</jats:italic>‐driven synthesis, mutagenesis, and radio‐tracer flux assays in HEK293 cells expressing the human DAT and SERT wild‐type and respective mutants. We found that only MCAT and the para‐CF<jats:sub>3</jats:sub>‐MCAT showed high selectivity, 200‐fold for DAT/SERT ‐ and 25‐fold for SERT/DAT, respectively. This suggests that the high selectivity achieved is determined rather by specific features of these compounds than by the volume of the para‐substituent. Accordingly, we were not able to find any correlation between the selectivity profile of the tested 4MCATs with either volume, polarity, or lipophilicity parameters. Next, based on our results, we tested the hypothesis that the two amino acids in SERT and DAT do not suffice to explain the selectivity between SERT and DAT. We have tested the para‐substituted methcathinones in the swapping mutations DAT Ser149Ala and SERT Ala169Ser and in line with our hypothesis, these mutations did not revert the selectivity profile found in the wild‐type transporters. Our findings rather suggest that the SERT selectivity achieved by the introduction of the CF<jats:sub>3</jats:sub>‐group in the para‐position of MCAT is not dependent on the volume of the para‐substituent but on specific chemical features of the fluorine atoms which may influence the on and off rate of the MCAT molecule on DAT and SERT.</jats:p><jats:p>This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in <jats:italic>The FASEB Journal</jats:italic>.</jats:p>