Discovery of Novel and Potent Small-Molecule Inhibitors of NO and Cytokine Production as Antisepsis Agents:  Synthesis and Biological Activity of Alkyl 6-(N-Substituted sulfamoyl)cyclohex-1-ene-1-carboxylate

Abstract

To develop a new therapeutic agent for sepsis, screening of the Takeda chemical library was carried out using mouse macrophages stimulated with lipopolysaccharide (LPS) to identify a new class of small-molecule inhibitors of inflammatory mediator production. The lead compound 5a was discovered, from which a series of novel cyclohexene derivatives I bearing a sulfamoyl and ester group were designed, synthesized and tested for their inhibitory activity against nitric oxide (NO) production. Derivatives I were synthesized by the coupling of sulfonyl chlorides and anilines with concomitant double bond migration in the presence of triethylamine, and phenyl ring substitution and modification of the ester and cyclohexene moieties were carried out. Among the compounds synthesized, ethyl (6R)-6-[N-(2-chloro-4-fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate [(R)-(+)-5n, TAK-242] was found to exhibit the most potent suppressive activity for the production of not only NO but also inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) induced by LPS-stimulated mouse macrophages with IC50 values of 1.8, 1.9 and 1.3 nM, respectively. It shows marked beneficial effects in vivo also. Intravenous administration of (R)-(+)-5n at doses of 0.1 mg/kg or more suppressed the production of NO and various cytokines [TNF-alpha, IL-6 and IL-1beta] in the mouse endotoxin shock model. Furthermore, it protected mice from death dose-dependently and all mice survived at a dose of 3 mg/kg. The minimum effective dose to protect mice from lethality in this model was 0.3 mg/kg, which was consistent with those for inhibitory effects on the production of NO and cytokines. Compound (R)-(+)-5n is currently undergoing clinical trials for the treatment of sepsis.

No Longer an Innocent Bystander: Epithelial Toll-Like Receptor Signaling in the Development of Mucosal Inflammation

“…The synthetic peptide-mimetic compound ST2825 prevents MyD88 homodimer formation leading to an inhibition in MyD88-dependent signaling, and prevents TLR9-dependent inflammation in vivo (120). A cyclohexene derivative, TAK-242, prevents TLR4 activation and was found to reduce the cytokine response to endotoxemic shock in mice (121,122). The Vaccinia virus protein A52R also reduces the TLR-mediated response by interacting with IRAK2 and TRAF6, and was found to reduce the cytokine response in an animal model of infectious otitis media (123,124) and to increase survival in models of endotoxemia (125).…”

Differences in the Pharmacokinetics of 4-Amino-3-Chlorophenyl Hydrogen Sulfate, a Metabolite of Resatorvid, in Rats and Dogs

“…Ethyl (6R)-6-[N-(2-chloro-4-fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate (CAS 243984-11-4, TAK-242, resatorvid) was discovered by Takeda Pharmaceutical Company Limited (Osaka, Japan) and was studied in clinical trials conducted in Japan, in the European Union, and in the United States as a potential new antisepsis drug, which acts as a cytokine production inhibitor (Yamada et al, 2005). Sepsis is a clinical condition defined by the presence of both infection (usually bacterial) and a systemic inflammatory response (Parrillo, 1993;Levy et al, 2003).…”

Optically Active Cyclohexene Derivative as a New Antisepsis Agent: An Efficient Synthesis of Ethyl (6R)-6-[N-(2-Chloro-4-fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate (TAK-242)

“…5) TAK-242 exhibited potent inhibitory activity for the production of not only nitric oxide (NO) but also various cytokines such as tumor necrosis factor-alfa (TNF-a), interleukin-1-beta (IL-1b) and IL-6 in vitro, as well as potent protective effects in the mouse endotoxin shock model. (R)-1 is currently undergoing clinical trails and is expected to be a promising antisepsis drug.…”

“…5) Although separation using preparative HPLC is a straightforward method for the preparation of small quantities of optically active compounds, applications to large scale production are sometimes troublesome. Thus, an efficient and practical process for the preparation of optically active (R)-1 was required in order to supply sufficient quantities for further evaluation of this compound.…”

“…12) Both products (R)-1 obtained via the diastereomeric esters, (6R,1ЈS)-6 and (6R,1ЈR)-7, were identical to an authentic sample prepared by HPLC separation. 5) As described above, a high yielding synthetic route for (R)-1 in high enantiomeric purity was established via the separation of diastereomeric ester (6RS,1ЈR)-7. This procedure, however, have remained unattractive due to annoying column chromatography and availability of the optically active alcohol (S)-5.…”