Biomimetic synthesis of hemiketal eicosanoids for biological testing

Abstract

The hemiketal (HK) eicosanoids HKE2 and HKD2 are the major products resulting from the biosynthetic cross-over of the 5-lipoxygenase and cyclooxygenase-2 pathways. They are formed by activated human leukocytes ex vivo, and, therefore, may be involved in regulation of the inflammatory response as autocrine or paracrine mediators. HKE2 and HKD2 are not commercially available and, so far, no method for their total chemical synthesis has been reported. The limited availability has impeded the characterization of their biological effects. Here, we describe a method for biomimetic preparation of HKE2 and HKD2 by reaction of recombinant human cyclooxygenase-2 with chemically synthesized 5S-HETE. We found that HKE2 did not induce or inhibit the release of TNFα and IL-1β by human THP-1 monocytes and phorbol ester treatment-derived macrophages.

Inhibition of 5‐Lipoxygenase‐Derived Leukotrienes and Hemiketals as a Novel Anti‐Inflammatory Mechanism of Urolithins

“…[ 7,8 ] The reaction of 5 S ‐HETE with COX‐2 generates an unstable di‐endoperoxide, an intermediate that rearranges into the biologically active HK eicosanoids, HKE 2 , and HKD 2 . [ 9 ] Their effect on angiogenesis, including the promotion of endothelial cell migration and tubulogenesis, [ 6 ] could have importance in IBD since the gut microvascular endothelium is a critical element in the onset and perpetuation of inflammation. [ 37 ] Thus, inhibition of HKE 2 and HKD 2 biosynthesis might be an attractive therapeutic antiangiogenic strategy by targeting neovascularization in IBD.…”

“…5 S ‐HETE can serve as a substrate of COX‐2, establishing a link between the 5‐LOX and COX‐2 pathways. The reaction of 5 S ‐HETE with COX‐2 generates an unstable di‐endoperoxide (equivalent to PGH 2 ), which is subsequently transformed to two hemiketal (HK) eicosanoids, HKE 2 and HKD 2 [ 6–9 ] ( Figure 1 ). PGs and LTs are well‐investigated molecules in IBD.…”

Roles of 5‐lipoxygenase and cyclooxygenase‐2 in the biosynthesis of hemiketals E2and D2by activated human leukocytes

“…5 S ‐HETE (15 μg) was added to 1 ml 100 mM Tris‐HCl buffer (pH 8) containing COX‐2 (25 nM), hematin (1 μM), and phenol (0.5 mM), and the reaction was incubated at 37°C for the first 5 min and then held at room temperature for 45 min (27). The sample was acidified to pH 3 with 1 N HCl, spiked with 50 μl methanol, and loaded on a 30‐mg HLB cartridge (Waters, Milford, MA, USA).…”

Total Synthesis and Biological Activity of the Arachidonic Acid Metabolite Hemiketal E2

“…6 Further study of HKD 2 and HKE 2 requires their total synthesis, as currently hemiketals D 2 /E 2 are produced enzymatically in small quantities starting from 5 S -HETE and recombinant COX-2. 7 Described herein is the first total synthesis of hemiketal E 2 ( 3 ).…”

“…Saponification of ester 21 provided hemiketal E 2 ( 3 ), identical by 1 H NMR to HKE 2 derived enzymatically. 7…”