α-Ketoheterocycles Able to Inhibit the Generation of Prostaglandin E2 (PGE2) in Rat Mesangial Cells

Abstract

Prostaglandin E₂ (PGE₂) is a key mediator of inflammation, and consequently huge efforts have been devoted to the development of novel agents able to regulate its formation. In this work, we present the synthesis of various α-ketoheterocycles and a study of their ability to inhibit the formation of PGE₂ at a cellular level. A series of α-ketobenzothiazoles, α-ketobenzoxazoles, α-ketobenzimidazoles, and α-keto-1,2,4-oxadiazoles were synthesized and chemically characterized. Evaluation of their ability to suppress the generation of PGE₂ in interleukin-1β plus forskolin-stimulated mesangial cells led to the identification of one α-ketobenzothiazole (GK181) and one α-ketobenzoxazole (GK491), which are able to suppress the PGE₂ generation at a nanomolar level.

Keywords: anti-inflammatory; inhibition; mesangial cells; prostaglandin E2; α-ketobenzothiazoles.

Figure 1

Generation of PGE₂ through metabolism of arachidonic acid.

Figure 2

Examples of α-ketothiazoles exhibiting anti-inflammatory properties.

Figure 3

Synthesis of α-ketobenzothiazoles 8a–h. (a) HN(OMe)Me·HCl, WSCI·HCl, DMAP, NMM, CH₂Cl₂; (b) (i) HN(OMe)Me·HCl, dry THF, −20 °C; (ii) i-PrMgCl, -20 °C; and (c) (i) n-BuLi, dry Et₂O, −78 °C; (ii) Weinreb amides 4a–h, dry Et₂O, −78 °C to rt.

Figure 4

Synthesis of compound 3d. (a) BrCH₂COOC(CH₃)₃, Bu₄NHSO₄, 50% NaOH, toluene; (b) 50% TFA, dry CH₂Cl₂.

Figure 5

Synthesis of compound 12. (a) Benzothiazole, n-BuLi, dry Et₂O, −78 °C to rt.

Figure 6

Synthesis of α-ketobenzoxazoles 17a,c,d and α-ketobenzimidazoles 17b,e. (a) (i) NaHSO₃, CH₂Cl₂; (ii) KCN, H₂O; (b) (i) CH₃COCl, CHCl₃/absolute EtOH; (ii) 2-aminophenol or 2-phenylenediamine, absolute EtOH; (c) Dess–Martin periodinane, CH₂Cl₂.

Figure 7

Synthesis of α-keto-1,2,4-oxadiazoles 22a–c. (a) TBDMSCN, KCN, 18-crown-6, dry CH₂Cl₂; (b) 50% aq. NH₂OH, microwave irradiation 50 W, 120 °C; (c) pivalic acid (for pivalate group) or benzoic acid (for benzoate group) or isobutyric anhydride (for isobutyrate group), DCC, dry CH₂Cl₂; (d) TBAF, toluene, microwave irradiation 90 W, 120 °C; (e) Dess–Martin periodinane, CH₂Cl₂.

Figure 8

Effect of compounds (A) GK181, (B) GK299, and (C) GK491 on IL-1/Fk-stimulated PGE₂ formation in mesangial cells. Cells were pretreated for 20 min with the indicated concentrations of GK compounds and then stimulated for 24 h in the absence (−) or presence (+) of 1 nM interleukin 1β(IL-1) plus 5 µM forskolin (Fk). Supernatants were taken for PGE₂ quantification using an enzyme-linked immunoassay as described in the Methods section. Data are presented as % of maximal IL-1/Fk stimulation and are means S.D. (n = 3). *** p < 0.001 considered statistically significant when compared to the unstimulated samples; ^# p < 0.05, ^## p < 0.01, ^### p < 0.001 compared to the IL-1/Fk-stimulated samples.

Figure 9

Proposed binding modes of GK181 (left) and GK491 (right) in the active site of GIIA sPLA₂ (PDB:1KQU). The purple ball is Ca²⁺, which coordinates with Asp48 and the glycine loop (Gly29, Gly31). The inhibitors are involved in π–π interactions with His6 and the catalytic His47.