Structure-activity relationships for substrate recognition by the human dopamine transporter

Information is available on the structure-activity relationships for dopamine as a substrate for uptake by the dopamine transporter. However, dopamine transport is a complex process involving substrate binding, translocation, release as well as transporter reorientation. The present study examines only the substrate recognition step by assessment of the potency of various dopamine-related compounds in inhibiting the binding of the cocaine analog [H-3]2beta-carbomethoxy-3beta-(4-fluorophenyl)tropane ([H-3]WIN 35,428) to human dopamine transporters expressed in HEK-293 cells. alpha-Methylation of the side chain, the presence of the amine, and the 2-carbon-length of the side chain were found to be important for binding affinity, whereas beta-hydroxylation of the side chain and methoxylation at the phenyl ring generated weaker compounds. In addition, the presence of both m- and p-OH at the phenyl ring bestowed an increase in potency but the presence of p-OH alone a decrease. N-alkylation (propylation or methylation) had little or an even slightly beneficial effect on affinity, whereas alpha-carbonylation and alpha-methanoylation reduced affinity. Amino naphthalene compounds with a fused benzenoid ring system retained some potency consonant with the extended (i.e. beta-rotameric) trans (=anti) form of the side chain in dopamine when interacting with the transporter. In a second series of experiments, the interaction between dopamine and structural variants was assessed by monitoring the capability of a compound to shift the dopamine inhibition curve to the right as expected for a competitive inhibitor acting at the same site. Appreciable deviation from competitive interaction was observed by removal of the amine from the side chain, by alpha-carbonylation, and by alpha-methanoylation. Two blocker-type compounds, semi-rigid variants of cocaine, also displayed significant deviation. A substrate-based compound, inhibiting cocaine analog binding without interfering with dopamine recognition, could be a cocaine antagonist allowing conformational changes to occur during dopamine uptake. (C) 2003 Elsevier Inc. All rights reserved.