Stereoisomers of an Aryl Pyrazole Glucocorticoid Receptor Agonist Scaffold Elicit Differing Anti-inflammatory Responses

Abstract

Glucocorticoids (GCs) are heavily prescribed to control inflammation in various human diseases; however, side effects associated with GCs are well documented and lead to serious metabolic and immunological complications with long-term use. The paradigm for GC function includes two well described modes of activity: dimer formation of the glucocorticoid receptor (GR) promotes transactivation, while monomeric interaction with co-regulators promotes transrepression. Previously, a set of aryl pyrazole-derived glucocorticoid receptor agonists (APGRAs) with potency rivaling current commercially available glucocorticoids were described. In this study, a further series of existing and novel stereopure APGRAs were thoroughly examined for biological activity and evaluated for structure-activity relationships (SARs). The si isomers with an upward OH moiety were ∼70% more active on average than the re isomers. Additionally, AP13 was found to elicit 79% transrepression of dexamethasone while eliciting less than half the transactivation response in 832/13 cells, a rat insulinoma cell line.

Scheme 1. Synthetic Route to APGRAs

Reagents and conditions: (ix) THF, −78 °C, 1 h.

Figure 1

¹H–¹³C HSQC NMR of AP13 (in CDCl₃) with assigned proton shifts. Y axis (f1) displays carbon signals from 0 to 160 ppm and X axis (f2) displays proton signals from 0 to 8 ppm.

Figure 2

¹H–¹H NOESY NMR of AP13 (in CDCl₃) with calculated distances between protons displayed on the left and their corresponding correlations boxed in red on the spectra to the right. Y axis (f1) and X axis (f2) displays proton signals from 0 to 4 ppm.

Figure 3

Biological analysis of top performing compounds using GR agonist and anti-inflammatory activity assays. (A) Structures of tested compounds. (B) CCL2-promoter-luciferase-reporter activity assay; x-axis, log concentration (molar); y-axis, %-maximal IL-1β response (relative promoter activity). (C) 3xGRE-promoter-luciferase-reporter activity assay; x-axis, log concentration (molar); y-axis, fold over control.

Figure 4

Top binding poses of AP13 (shown in blue) and AP14 (shown in magenta) superimposed in the LBP of the glucocorticoid receptor 3BQD (shown in green). Helices labeled in black as H1 through H12.