Identification of Novel Cyclooxygenase-1 Selective Inhibitors of Thiadiazole-Based Scaffold as Potent Anti-Inflammatory Agents with Safety Gastric and Cytotoxic Profile

Abstract

Major obstacles faced by the use of nonsteroidal anti-inflammatory drugs (NSAID) are their gastrointestinal toxicity induced by non-selective inhibition of both cyclooxygenases (COX) 1 and 2 and their cardiotoxicity associated with a certain class of COX-2 selective inhibitors. Recent studies have demonstrated that selective COX-1 and COX-2 inhibition generates compounds with no gastric damage. The aim of the current study is to develop novel anti-inflammatory agents with a better gastric profile. In our previous paper, we investigated the anti-inflammatory activity of 4-methylthiazole-based thiazolidinones. Thus, based on these observations, herein we report the evaluation of anti-inflammatory activity, drug action, ulcerogenicity and cytotoxicity of a series of 5-adamantylthiadiazole-based thiazolidinone derivatives. The in vivo anti-inflammatory activity revealed that the compounds possessed moderate to excellent anti-inflammatory activity. Four compounds 3, 4, 10 and 11 showed highest potency (62.0, 66.7, 55.8 and 60.0%, respectively), which was higher than the control drug indomethacin (47.0%). To determine their possible mode of action, the enzymatic assay was conducted against COX-1, COX-2 and LOX. The biological results demonstrated that these compounds are effective COX-1 inhibitors. Thus, the IC₅₀ values of the three most active compounds 3, 4 and 14 as COX-1 inhibitors were 1.08, 1.12 and 9.62 μΜ, respectively, compared to ibuprofen (12.7 μΜ) and naproxen (40.10 μΜ) used as control drugs. Moreover, the ulcerogenic effect of the best compounds 3, 4 and 14 were evaluated and revealed that no gastric damage was observed. Furthermore, compounds were found to be nontoxic. A molecular modeling study provided molecular insight to rationalize the COX selectivity. In summary, we discovered a novel class of selective COX-1 inhibitors that could be effectively used as potential anti-inflammatory agents.

Keywords: anti-inflammatory; cyclooxygenase; cytotoxicity; enzyme inhibition; lipoxygenase; molecular modeling; thiadiazole; ulcerogenic effect.

Figure 1

Representative COX-1 selective inhibitors with improved safety profile.

Figure 2

5-methylthiazole-based thiazolidinones and 5-adamantylthiadiazole-based thiazolidinones.

Scheme 1

Preparation of final derivatives. Synthesis of title compounds. Reagents and conditions: (a) conc. H₂SO₄, 15 min 0 °C, r.t. 30 min; (b) NH₃; (c) DMF, 3 h, r.t.; (d) NH₄SCN, ethanol, reflux, 1 h, r.t overnight; (e) CH₃COOH/CH₃COONa, reflux, 3–5 h.

Figure 3

Three- and two-dimensional representations of the binding mode of ibuprofen and compounds 3, 4 and 14 in COX-1 active site (PDB 1EQG). H-bond, halogen, pi–pi, pi–alkyl and pi-cation interactions are represented as dotted lines in green, blue, violet, grey and orange, respectively.

Figure 3

Three- and two-dimensional representations of the binding mode of ibuprofen and compounds 3, 4 and 14 in COX-1 active site (PDB 1EQG). H-bond, halogen, pi–pi, pi–alkyl and pi-cation interactions are represented as dotted lines in green, blue, violet, grey and orange, respectively.

Figure 4

Superimposed COX-2 binding sites from the two co-crystals PDB 3LN1 and PDB 6BL3 showing Arg120 conformation alteration.

Figure 5

Three- and two-dimensional representations of the binding mode of compound 3 in COX-2 active site (PDB 3LN1 and 6BL3). H-bond, halogen, pi–pi, pi–alkyl and pi-cation interactions are represented as dotted lines in green, blue, violet, grey and orange, respectively.

Figure 5

Three- and two-dimensional representations of the binding mode of compound 3 in COX-2 active site (PDB 3LN1 and 6BL3). H-bond, halogen, pi–pi, pi–alkyl and pi-cation interactions are represented as dotted lines in green, blue, violet, grey and orange, respectively.

Figure 6

(A) Two-dimensional interaction diagram of the reference compound NDGA in 5-LOX. (B) Two-dimensional interaction diagram of compound 14 in 5-LOX. (C) Superposition of compound 14 (blue) bound to 5-LOX by comparison with NDGA (magenta), with labeled specific residues. Active site zinc revealed as yellow small sphere, green and red dotted arrows illustrate hydrogen bond, pink dotted arrows represent halogen bond and yellow sphere hydrophobic interactions.

Figure 7

Cell proliferation assessment of MRC-5 cell lines subjected to different derivatives in cell culture. MRC-5 cell lines grown in culture were incubated separately with each of the derivatives at concentrations of 0.1, 1 and 10 μM for 48 h. The cell number was analyzed in cell cultures with microscopic techniques (Neubauer counting chamber), as reported in “Methods” and expressed as % of the proliferation of cultures involving control untreated cells. The studies displayed above show the mean numbers ± SD of two independent biological tests (n ≥ 3). The diagram illustrated above and the t-test statistical analysis were performed utilizing the GraphPad Prism 6.0 program. No significant statistical difference between the control untreated culture cells with each tested derivative was attained.