Exploring 6-Azaindole and 7-Azaindole Rings for Developing Cannabinoid Receptor 1 Allosteric Modulators

Abstract

Introduction and Objective: Org27569 is a prototypical allosteric modulator of the cannabinoid receptor 1 (CB₁). It belongs to the indole-2-carboxamide scaffold and has been intensively investigated in pharmacology and in structure-activity relationship (SAR) studies. Although azaindoles are rare in natural products and differ only by the presence of an extra ring nitrogen, they were demonstrated as valuable bioisosteres in many pharmacologically important molecules. To extend the SAR investigation of the indole-2-carboxamide class of CB₁ allosteric modulators, azaindole (pyrrolopyridine) rings were used to replace the indole ring of Org27569 analogs to explore the potential of azaindole-2-carboxamides as CB₁ allosteric modulators. Using 6- and 7-azaindole in lieu of the indole moiety within this class of CB₁ allosteric modulators indeed improved the aqueous solubility. Materials and Methods: We synthesized 6- and 7-azaindole-2-carboxamides and their indole-2-carboxamide counterparts. The molecules were evaluated by [³H]CP55,940 binding and [³⁵S]GTPγS binding assays for their allosteric modulation of the CB₁ receptor. Results: The 7-azaindole-2-carboxamides lost the ability to bind to the CB₁ receptor. The 6-azaindole-2-carboxamides (e.g., 3c and 3d) showed markedly reduced binding affinities to the CB₁ receptor in comparison with their indole-2-carboxamide counterparts. However, they behaved similarly as indole-2-carboxamides in potentiating the orthosteric agonist binding and inhibiting the orthosteric agonist-induced G-protein coupling. The results indicated that some azaindole scaffolds (e.g., 6-azaindole) are worth further exploration, whereas the 7-azaindole ring is not a viable bioisostere of the indole ring in the Org27569 class of CB₁ allosteric modulators.

Keywords: CB1 receptor; allosteric modulators; azaindole; bioisostere; cannabinoid.

FIG. 1.

Prototypical CB₁ allosteric modulator Org27569 (A), referenced indole-2-carboxamide analogs of Org27569 (B), and designed 6- and 7-azaindole-2-carboxamides (C). CB₁, cannabinoid receptor 1.

FIG. 2.

Synthesis of indole- and azaindole-2-carboxamides. DMTMM, 4-(4,6-Dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride; NMM, N-methyl morpholine; THF, tetrahydrofuran.

FIG. 3.

Response of 3c (LDK1314) and 3 d (LDK1316) on [³⁵S]GTPγS binding to membranes expressing CB₁. The effects of 0.1 μM CP55,940 alone, or 0.1 μM CP55,940 with the allosteric modulators 3c or 3d, 3c, or 3d alone, or 1.0 μM SR141716A alone on [³⁵S]GTPγS binding were measured at the concentrations indicated. Data are presented as a percentage of basal levels of [³⁵S]GTPγS binding. Nonspecific binding was measured in the presence of 10 μM unlabeled GTPγS. Each data point represents the mean±standard error of the mean of at least three independent experiments performed in duplicate. GTPγS, guanosine 5′-O-[gamma-thio]triphosphate.