Fig. 4: Orthogonal validation and profiling of 36.
From: Development of Nurr1 agonists from amodiaquine by scaffold hopping and fragment growing
a Effects of 36 on the human full-length Nurr1 homodimer (NurRE) and monomer (NBRE). Data are the mean±S.E.M. fold activation vs. DMSO ctrl; n ≥ 3. b Effects of 36 on the Nurr1-RXR heterodimer (DR5) in the absence and presence of bexarotene (0.1 µM). Data are the mean±S.E.M. fold activation vs. DMSO ctrl or vs. 0.1 µM BEX; n ≥ 3. c Binding of 36 to the Nurr1 LBD (Kd 0.17 µM, n = 1.0) determined by ITC. The fitting of the heat of binding is shown and the isotherm at 25 °C is shown as inset. d Effects of 36 and 29 on the expression of the Nurr1-regulated tyrosine hydroxylase (TH), vesicular amino acid transporter 2 (VMAT2), and superoxide dismutase 2 (SOD2) in astrocytes (T98G). Data are the mean±S.E.M. fold mRNA induction vs. DMSO control; n = 3; #p < 0.1, *p < 0.05 (t-test vs. DMSO ctrl). e Selectivity screening of 36 on nuclear receptors. Heatmap shows the mean relative activation compared to reference ligands (listed in the methods section); n = 3. (f) 36 (10 µM) had no toxic effect in a WST-8 assay in N27 rat neurons and HEK293T cells. AQ (1, 10 and 30 µM) was toxic. Data are the mean±S.E.M. rel. absorbance (450 nm); n ≥ 3. g Permeability of Nurr1 agonists in a parallel artificial membrane permeability assay (PAMPA) and in a cellular model of the blood-brain-barrier (BBB). Propranolol and the brain-penetrant reference antipyrine for comparison. Data are the mean ± SD; n = 6. h Metabolism of 36 by rat liver microsomes resulted in demethylation of the dimethylamino group. Data are the mean ± SD; n = 4.
