PT - JOURNAL ARTICLE AU - Richard B. Rothman AU - Subramaniam Ananthan AU - John S Partilla AU - Surendra K Saini AU - Omar Moukha-Chafiq AU - Vibha Pathak AU - Michael H Baumann TI - Studies of the Biogenic Amine Transporters. 15. Identification of Novel Allosteric Dopamine Transporter Ligands with Nanomolar Potency AID - 10.1124/jpet.114.222299 DP - 2015 Jan 01 TA - Journal of Pharmacology and Experimental Therapeutics PG - jpet.114.222299 4099 - http://jpet.aspetjournals.org/content/early/2015/03/18/jpet.114.222299.short 4100 - http://jpet.aspetjournals.org/content/early/2015/03/18/jpet.114.222299.full AB - Abstract.Novel allosteric modulators of the dopamine transporter (DAT) have been identified. For example, we have shown previously that N-(diphenylmethyl)-2-phenyl-4-quinazolinamine (SRI-9804), N-(2,2-diphenylethyl)-2-phenyl-4-quinazolinamine (SRI-20040), and N-(3,3-Diphenylpropyl)-2-phenyl-4-quinazolinamine (SRI-20041) partially inhibit [125I]RTI-55 binding and [3H]dopamine ([3H]DA) uptake, slow the dissociation rate of [125I]RTI-55 from the DAT, and allosterically modulate d-amphetamine-induced DAT-mediated dopamine (DA) release. Over 500 analogs of these ligands have been synthesized and evaluated for activity as allosteric modulators of DAT. We report here on 36 selected compounds. Using synaptosomes prepared from rat caudate, we conducted [3H]DA uptake inhibition assays, DAT binding assays with [3H]WIN35428, and DAT-mediated release assays with either [3H]MPP+ or [3H]DA. The initial set of compounds binned into three groups of [3H]DA uptake inhibitors: 1) full efficacy agents with a one-site fit, 2) full efficacy agents with a two-site fit and 3) partial efficacy agents with a one-site fit. We focused further studies on the partial efficacy uptake inhibitors. These agents were partial inhibitors of DA, serotonin, and norepinephrine uptake, yet much less potent at inhibiting [3H]WIN35428 binding to DAT. For example, SRI-29574 partially inhibited uptake at DAT with an IC50 = 2.3±0.4 nM, without affecting binding to DAT. Overall, at concentrations less than 1 µM, these agents did not alter DAT-mediated release of [3H]MPP+ in the absence or presence of 100 nM d-amphetamine. At a dose 25-times greater than its IC50 for DAT uptake inhibition, SRI-29574 had no significant effect on the d-amphetamine EC50 or Emax value for DAT-mediated release of [3H]MPP+. These studies demonstrate the existence of potent DAT ligands that partially block [3H]DA uptake, without affecting DAT binding or d-amphetamine-induced [3H]MPP+ release. These compounds may prove to be useful probes of biogenic amine transporter function as well as novel therapeutics.