Fig. 3. Identification of high-affinity allosteric inhibitors for SERT.

a Chemical structure of three tested compounds, all based on S-CIT template with an amide instead of the cyano in S-CIT. They differ in having either none (AE), one (Lu AF60097) or two (AF) double bonds (stippled circles) in their N-substituent. b The allosteric potency of the compounds inhibiting the dissociation of [³H]S-CIT is within the micromolar range: Lu AF60097 (red), IC₅₀ = 6.50 [5.07; 8.31] µM; AF (brown), IC₅₀ = 10.4 [8.45; 12.9] µM; and AE (black), IC₅₀ = 33.8 [30.7; 37.3] µM. Data are mean [S.E. interval], n = 3-5. c The allosteric potency increases to nanomolar concentrations when assessed by inhibition of [³H]IMI dissociation. IC₅₀ values (in nM) for Lu AF60097, AF and AE are 31.4 [25.2; 39.1], 192 [173; 215], and 119 [103; 138], respectively (mean [S.E. interval], n = 3–6. d A zoomed-in view of S1:IMI (in purple)/S2:Lu AF60097 (in green), and e S1:S-CIT (in salmon)/S2:Lu AF60097 (in green) conditions. f Distribution of the Thr497 χ1 rotamer for S1:S-CIT/S2:Lu AF60097 (salmon) and S1:IMI/S2:Lu AF60097 (purple) conditions. g Distribution of the Glu494/S2:Lu AF60097 distance (minimum distance between the charged N of Lu AF60097 and the two carboxyl oxygens of Glu494) for S1:S-CIT/S2:Lu AF60097 (salmon) and S1:IMI/S2:Lu AF60097 (purple) conditions. Experiments in b and c are performed essentially as in Fig. 1 on membrane preparations from COS-7 cells transiently transfected with SERT WT. Data are shown as means ± SEM (error bars). Source data are provided as a Source Data file.