Study of Chalcogen Aspirin Derivatives with Carbonic Anhydrase Inhibitory Properties for Treating Inflammatory Pain

Abstract

Carbonic anhydrase (CA) inhibitors represent intriguing tools for treating pain. This study aims at studying the pharmacological profile of chalcogen bioisosteres of aspirin, as inhibitors of CA isoforms (hCA I, II, IV, VII, IX, and XII). Our results show that selenoaspirin (5) displayed markedly superior inhibitory potency across all tested isoforms compared to thioaspirin (7) and aspirin, with a strong selectivity against the isoform CA IX. X-ray crystallography confirmed that both compounds bind effectively within the active site of hCA II, revealing unique structural characteristics compared to those of aspirin. In a preclinical model of inflammatory pain, compound 7 exhibited a longer lasting antihyperalgesic effect than aspirin, though with a lower potency. Conversely, compound 5 exhibited both lower potency and efficacy than aspirin in reducing pain, which entailed both adverse effects. Nevertheless, the therapeutic potential of chalcogen-based aspirin derivatives as novel CA inhibitors deserves to be further explored for clinical applications.

Figure 1

Molecular structure of aspirine and its sulfur- and selenium-containing analogues.

Scheme 1. Synthetic Route for the Synthesis of Selenoaspirine 5 from Anthranilic Acid 1

Scheme 2. Synthesis of Thioaspirine 7 from Thiosalicylic Acid 6

Figure 2

A) Active site region of hCA II/hydrolyzed form of 5 adduct (PDB: 9FAI). Inhibitor is labeled in green, van der Waals interactions in blue, and water bridges as well as hydrogen bonds in red. B) Active site region of hCA II/hydrolyzed form of 7 adduct (PDB: 9FAO). Inhibitor is labeled in cyan, van der Waals interactions in blue, and water bridges as well as hydrogen bonds in red.

Figure 3

A) Overlay of the hydrolyzed form of selenoaspirin 5 (green) and thioaspirin 7 (cyan) with hCA II. B) Overlay of the hydrolyzed form of aspirin (magenta, PDB: 6UX1), selenoaspirin 5 (green), and thioaspirin 7 (cyan) with hCA II. Specific residues are labeled.

Figure 4

Effect of acute administration of aspirin and compounds 5 and 7 on thermal hyperalgesia associated with inflammatory pain in CFA-injected mice. Inflammatory pain was induced by intraplantar injection of CFA. Control animals were treated with vehicles. Thermal hyperalgesia was evaluated by Hot Plate test, pain-related behavior (i.e., licking of the hind paw) was observed, and the time (seconds) of the first sign was recorded. Twenty-four hours after CFA injection, aspirin (10–200 mg kg^–1; MW 180.16; A), 7 (220–330 mg kg^–1; MW 196.22; B), and 5 (270–405 mg kg^–1; MW 243.13; C) were orally administered and measurements assessed before and every 20 min after injection. ^∧∧p < 0.01 vs CTR + vehicle treated animals. *p < 0.05 and **p < 0.01 vs pretest (0 min). Each value represents the mean of 6 mice performed in 2 different experimental sets.

Figure 5

Effect of acute administration of aspirin and compounds 5 and 7 on mechanical hyperalgesia associated with inflammatory pain in CFA-injected mice. Inflammatory pain was induced by the intraplantar injection of CFA. Control animals were treated with vehicles. Mechanical hyperalgesia was evaluated by the Paw Pressure test, and data are expressed as the latency of animal paw withdrawal (s). Twenty-four hours after CFA injection, aspirin (10–200 mg kg^–1; MW 180.16; A), 7 (220–330 mg kg^–1; MW 196.22; B), and 5 (270–405 mg kg^–1; MW 243.13; C) were orally administered and measurements assessed before and every 20 min after injection. ^∧∧p < 0.01 vs CTR + vehicle treated animals. *p < 0.05 and **p < 0.01 vs pretest (0 min). Each value represents the mean of 6 mice performed in 2 different experimental sets.

Figure 6

Effect of acute administration of aspirin, 5, and 7 on mechanical allodynia associated with inflammatory pain in CFA-injected mice. Inflammatory pain was induced by the intraplantar injection of CFA. Control animals were treated with vehicles. Mechanical allodynia was evaluated by the Von Frey test, and data are expressed as the weight (g) that the animal tolerated on the paw. Twenty-four hours after CFA injection, aspirin (50–200 mg kg^–1; MW 180.16; A), 7 (220–330 mg kg^–1; MW 196.22; B), and 5 (270 mg kg^–1; MW 243.13; C) were orally administered and measurements assessed before and every 20 min after injection. ^∧∧p < 0.01 vs CTR + vehicle treated animals. *p < 0.05 and **p < 0.01 vs pretest (0 min). Each value represents the mean of 6 mice performed in 2 different experimental sets.

Figure 7

Effect of aspirin and compound 7 on trans-epithelial permeability. FITC-dextran fluorescence intensity measured in Caco-2 cells treated with aspirin (10 mM) or compound 7 (10 mM and 16.5 mM) for 72 h. Each column represents the mean ± SEM (n = 5 independent experiments). *P < 0.05 significant difference versus control cells (CTR).