Design, Synthesis, and Biological Evaluation of Some Novel Pyrrolizine Derivatives as COX Inhibitors with Anti-Inflammatory/Analgesic Activities and Low Ulcerogenic Liability

Abstract

Non-steroidal anti-inflammatory drugs (NSAIDs) are the most commonly prescribed anti-inflammatory and pain relief medications. However, their use is associated with many drawbacks, including mainly serious gastric and renal complications. In an attempt to circumvent these risks, a set of N-(4-bromophenyl)-7-cyano-6-substituted-H-pyrrolizine-5-carboxamide derivatives were designed, synthesized and evaluated as dual COX/5-LOX inhibitors. The structural elucidation, in vivo anti-inflammatory and analgesic activities using a carrageenan-induced rat paw edema model and hot plate assay, were performed, respectively. From the results obtained, it was found that the newly synthesized pyrrolizines exhibited IC50 values in the range of 2.45-5.69 µM and 0.85-3.44 µM for COX-1 and COX-2, respectively. Interestingly, compounds 12, 13, 16 and 17 showed higher anti-inflammatory and analgesic activities compared to ibuprofen. Among these derivatives, compounds 16 and 19 displayed better safety profile than ibuprofen in acute ulcerogenicity and histopathological studies. Furthermore, the docking studies revealed that compound 17 fits nicely into COX-1 and COX-2 binding sites with the highest binding affinity, while compound 16 exerted the highest binding affinity for 5-LOX. In light of these findings, these novel pyrrolizine-5-carboxamide derivatives represent a promising scaffold for further development into potential dual COX/5-LOX inhibitors with safer gastric profile.

Keywords: 5-LOX; COX; analgesic; anti-inflammatory; pyrrolizine; ulcerogenicity.

Figure 1

The IC₅₀ values of pyrrolizine-based anti-inflammatory agents 1–3 linked with a carboxylic acid moiety.

Figure 2

The IC₅₀ values of pyrrolizine-based anti-inflammatory agents 4–6 lacking the free carboxylic acid group.

Scheme 1

Synthesis of compound 12. Reagents and reaction conditions: (a) (CH₃)₂SO₄, benzene, CH₂(CN)₂; (b) ClCH₂COCl, AcOH, CH₃COONa; (c) acetone, K₂CO₃, reflux, 24 h.

Scheme 2

Synthesis of compounds 13–16. Reagents and reaction conditions: (a) ClCH₂COCl, benzene, 48 h; (b) 1-methylpiperazine, NaHCO₃, absolute ethanol, reflux, 6 h; (c) KHCO₃, DMF, rt, 48 h; (d) benzoyl chloride, benzene, rt, 48 h.

Scheme 3

Synthesis of compounds 17–19. Reagents and reaction conditions: (a) p-toluenesulfonyl chloride, acetone, K₂CO₃; rt, 12 h; (b) oxalyl chloride, dry acetone, rt, 24 h; (c) 1. ibuprofen, SOCl₂; 2. compound 12, dry benzene, rt, 24 h.

Figure 3

Design strategies for compounds 12–18.

Figure 4

Change in edema thickness using carrageenan-induced rat paw edema mode of compounds 12–19 at 1 h (${}^{▄}$), 2 h (${}^{▄}$) and 3 h (${}^{▄}$) after induction of inflammation; data expressed as mean ± SEM, (n = 6); data were analyzed by One way ANOVA followed by student-Newman-Keuls multiple comparison test; ^a statistically significant from control (p < 0.001); ^b statistically significant from control (p < 0.01); ^c statistically significant from control (p < 0.05).

Figure 5

Relationship between the in vivo anti-inflammatory (at 3 h) and analgesic (at 2 h) activities of the new compounds 12–19 at a dose of 0.48 mmol/kg with the modifications in the chemical structure; * activity means both anti-inflammatory and analgesic activities.

Figure 6

The analgesic activity results using hot-plate test for control, compound 12–19, and ibuprofen at 0.24 (${}^{▄}$) and 0.48 mmol/kg (${}^{▄}$); data were represented as means ± SEM, n = 6; data were analyzed by One way ANOVA followed by student-Newman-Keuls multiple comparison test; % change = 100 × (T₁ − T₀)/T₀; ^a statistically significant from control (p < 0.001); ^b statistically significant from control (p < 0.01); ^c statistically significant from control (p < 0.05); ^d statistically not significant from control.

Figure 7

The histological TSs (C–L) in the stomach of rat treated with compounds 12–19, respectively in comparison to control (A) and ibuprofen (B), using haematoxylin and eosin stain, 400×.

Figure 8

Ibuprofen docked superimposed onto the position of co-crystalized ligand within RMSD of 0.62 Å, and exhibited two hydrogen bonds between its COOH group and NH₂ of Arg120.

Figure 9

(A) Validation of the performance of AutoDock program by docking of the native co-crystallized ligands S58 into COX-2; (B) Validation of the performance of AutoDock program by docking of arachidonic acid into 5-LOX.

Figure 10

Docking mode of compound 17 (ball and stick) into COX-2 (pdb code: 1cx2). It revealed four hydrogen bonds (green dotted lines) with Arg120, Leu352, and Tyr355, within RMSD of 1.45 Å from the co-crystallized S58 ligand (yellow sticks).

Figure 11

The binding mode of the interaction of compound 19 (ball and stick) into the target enzyme (5-LOX; pdb code: 3o8y). It bounds with Tyr181 by one hydrogen bond (green dotted lines) within RMSD of 3.24 Å from the reference ACD (yellow sticks).

Figure 12

(A) The binding interactions of compound 18 into COX-1; (B) binding interactions of compound 18 into COX-2; and (C) binding interactions of compound 18 into 5-LOX illustrated by Ligplot.