Fig 1.
Structure of (S)-xamoterol and two sites of modification.
Fig 2.
Synthesis of (R)- and (S)-xamoterol.
Fig 3.
General method for the synthesis of compounds STD-101-B1 to B8.
Fig 4.
Synthesis of STD-101-B9.
Fig 5.
General method for the synthesis of compounds STD-101-D1 to D6.
Fig 6.
Syntheses of the amine components for STD-101-D5 and D6.
Fig 7.
Effects of structural modifications of the phenolic OH moiety of xamoterol on the cAMP pathway mediated by ADRB1.
clogPa, Calculated with ChemDraw Pro Version 16.0 (PerkinElmer Health Sciences, CT; EC50 (nM)b, Geometrical mean of EC50 values from at least two independent experiments; % Iso maxc, Percent efficacy compared to the maximum response achieved with isoproterenol; ~d, Could not be determined.
Fig 8.
Concentration-response effects of compounds on the cAMP pathway via ADRB1.
Data are expressed as a percentage of maximum efficacy obtained with the full agonist isoproterenol. Values represents means ± S.E.M.s (1–2 experiments with n = 1–2).
Fig 9.
Effects of structural modifications of the morpholino urea moiety of xamoterol on the cAMP pathway mediated by ADRB1.
clogPa, Calculated with ChemDraw Pro Version 16.0; EC50 (nM)b, Geometrical mean of EC50 values from at least two independent experiments; % Iso maxc, Percent efficacy compared to the maximum response achieved with isoproterenol; ~d, Could not be determined.
Fig 10.
Concentration-response effects of compounds on the β-arrestin pathway via ADRB1.
Data are expressed as a percentage of maximum efficacy obtained with the full agonist isoproterenol. Values represents means ± S.E.M.s (1–2 experiments with n = 1–2). Xamoterola; data has been previously published [11].
Fig 11.
Crystal structure of ADRB1 with xamoterol (S) (cyan, panel A), xamoterol (R) (blue, panel B), and STD-101-D1 (salmon, panel C) docked into the ligand-binding site. The transmembrane regions are shown as green ribbons, and. Putative interactions are displayed as yellow dashed lines with estimated distance in angstroms (Å). The carbon-nitrogen chain of xamoterol (S) is predicted to pack approximately 1.0 Angstroms closer to D121 and Transmembrane Helix 3 compared to xamoterol (R), and its morpholino ring rests in a rotated pose as well.
Fig 12.
Inhibitory effects of ADRB1 ligands on LPS-induced TNFα response in primary microglia.
Data are represented as mean ± S.E.M.s of four independent experiments (n = 3–18 per group, * p < 0.05, **p < 0.01, *** p < 0.001, one-way ANOVA followed by Dunnett’s multiple comparison against LPS exposure alone).
Fig 13.
Inhibitory effects of ADRB1 ligands on LPS-induced TNFα response in mice.
(A) Plasma TNFα concentrations in control animals and animals pretreated with xamoterol or STD-101-D1 90 min after LPS injection. (B) TNFα, IL1β, and IL6 mRNA expression in homogenized cortical tissue from control mice and animals pretreated with xamoterol or STD-101-D1 90 min after LPS injection. Data are represented as mean ± S.E.M.s of three independent experiments. (n = 4–14 per group, * p < 0.05, **p < 0.01, *** p < 0.001, one-way ANOVA followed by Dunnett’s multiple comparison).
Fig 14.
Metabolic stability in mouse, rat, and human microsomes.
STD-101-D1 and two reference compounds verapamil and propranolol were incubated at 0.1 uM in mouse (A), rat (B), or human (C) liver microsomes. Serial samples were removed up until 60 min. All experiments were performed in duplicate, and data are represented as mean ± S.E.M.
Fig 15.
Pharmacokinetics of xamoterol and STD-101-D1.
Systemic (A) and portal vein (B) plasma concentrations of xamoterol and STD-101-D1 as a function of time after a single injection of xamoterol (10 mg/kg) or STD-101-D1 (10 mg/kg) via intravenous (IV), intraperitoneal (IP) and oral (PO) administration. Plasma and brain (C) concentrations of xamoterol and STD-101-D1 in rats collected 20 min after a single injection of xamoterol or STD-101-D1 (10 mg/kg) via IV, IP, and PO administration. Data are represented as mean ± SEM (n = 3 per route).
Table 1.
Pharmacokinetic parameters of xamoterol and STD-101-D1 determined in the 4-hr time-course study.
Table 2.
Pharmacokinetic parameters of xamoterol and STD-101-D1 determined in the 20-min post-dose collection study.
Fig 16.
Effects of xamoterol and STD-101-D1 on heart rate and blood pressure.
Changes in heart rate (A) and blood pressure (B) following subcutaneous administration of xamoterol or STD-101-D1 at dose of 3 mg/kg. Date are represented as mean ± SEM (n = 3 per compound). (One-sample t-test vs. 0% theoretical mean, * p < 0.05).