C. A. Veale

Stereocontrolled total synthesis of lipoxins A

Synthese de la lipoxine A (acide trihydroxy-5,6,15 eicosatetraene-7,9,11,13oique) et de ses isomeres tout-trans

Actions of Lipoxin A4 and Related Compounds in Smooth Muscle Preparations and on the Microcirculation in Vivo

The present chapter summarizes our findings with lipoxins (LX) in spasmogenic assays and in the intact microvasculature of the hamster cheek pouch. The initial observations1,2 were made in experiments using lipoxins isolated from human leukocytes. With the aid of synthetic compounds3, it has been possible to further explore the pharmacodynamics and structure activity relationships for lipoxins in smooth muscle preparations.

Identification of a novel 7-cis-11-trans-lipoxin A4 generated by human neutrophils: total synthesis, spasmogenic activities and comparison with other geometric isomers of lipoxins A4 and B4

Addition of (15S)-hydroxy-5,8,11-cis-13-trans-eicosatetraenoic acid (15-HETE) and the ionophore A23187 (2.5 microM) to human neutrophils led to the formation of both lipoxin A4 and lipoxin B4 as well as a novel 5,6,15-trihydroxyeicosatetraenoic acid. The new compound was identified using an improved isolation and detection system and its basic structure was determined by physical methods. On the basis of biosynthetic considerations, geometric isomers of lipoxin A4 and lipoxin B4 were prepared by total synthesis. Comparison of these synthetic materials with the neutrophil-derived product showed that the new compound is (5S,6R,15S)-trihydroxy-9,11,13-trans-7-cis-eicosatetraenoic acid or the 7-cis-11-trans-isomer of LXA4 (7-cis-11-trans-LXA4). LXA4, 11-trans-LXA4, 7-cis-LXA4 and 7-cis-11-trans-LXA4 all evoked dose-dependent (0.1-10 microM) contractions of the guinea pig lung strip, whereas 6-cis-LXB4 and 6-cis-8-trans-LXB4 relaxed this preparation. LXA4 and 7-cis-LXA4 were approx. 10-times more potent than the compounds with 11-trans geometry. However, all four double-bond isomers of LXA4 caused contractions which, based upon pharmacological evidence, appeared to involve specific activation of the same site as cysteinyl-containing leukotrienes. In conclusion, 7-cis-11-trans-LXA4 was isolated and identified as a novel biologically active eicosanoid formed by human neutrophils.

Ca2+ mobilization with leukotriene A4 and epoxytetraenes in human neutrophils

The biosynthesis of leukotrienes and lipoxins involves epoxide-containing intermediates which may be subject to several routes of transcellular metabolism. We have examined the capacity of leukotriene A4 (LTA4) and 15S-hydroxy-5,6-oxido-7,9,13-trans-11-cis-eicosatetraenoic acid [5(6)-epoxytetraene] to stimulate the mobilization of free cytosolic calcium [( Ca2+]i) in human blood neutrophils. To gain insight into structure-activity relationships, a putative intermediate in lipoxin biosynthesis, 5S-hydroxy-14,15-oxido-6,10,12-trans-8-cis-eicosatetraenoic acid [14(15)-epoxytetraene], was prepared by total synthesis. When added to fura-2 loaded neutrophils, each of these compounds provoked a rapid and transient increase in [Ca2+]i (maximum by 8 sec) which returned to baseline within 60-90 sec. Ca2+ mobilization with LTA4 was dose dependent and, at 1 microM, the efficacies of LTA4 and LTB4 were quantitatively similar. The 5(6)-epoxytetraene and 14(15)-epoxytetraene were less potent than LTA4. Prior exposure of the cells to ethyleneglycolbis(aminoethylether)tetra-acetate (EGTA) (60 sec, 3 mM) did not diminish either the amplitude or the extent of [Ca2+]i elicited by LTA4. Methyl esters of LTA4 and 5(6)-epoxytetraene were less potent than their corresponding free acids, whereas the free acid of 14(15)-epoxytetraene and its methyl ester were quantitatively similar. Results from alcohol trapping studies showed that these epoxides were intact during the initial phase of Ca2+i mobilization (t0-10 sec) stimulated by LTA4, 5(6)-epoxytetraene, and 14(15)-epoxytetraene. In addition, the individual mixtures of products formed upon aqueous hydrolysis of each of the epoxides did not stimulate changes in [Ca2+]i. In each case, the products formed were identified by physical methods including reverse phase high pressure liquid chromatography, ultraviolet spectroscopy and gas liquid chromatography-mass spectrometry. These results indicate that, when added to human neutrophils, LTA4, 5(6)-epoxytetraene and 14(15)-epoxytetraene each stimulate a rapid mobilization of [Ca2+]i. Moreover, they suggest that intermediates in the biosynthesis of leukotrienes and lipoxins possess intrinsic activities that may serve to amplify cellular responses within their cell of origin or act on adjacent cells during their transcellular metabolism.

Pharmacodynamics of lipoxin A4 in airway smooth muscle

The lipoxins (LX) are a novel group of arachidonic acid metabolites recently discovered in human leukocytes [for reviews see 1]. The lipoxins have biological activities [2] which suggest a potential as mediators or modulators of inflammation. Thus, LXA 4 (5S, 6R, 15S-trihydroxy-7,9,13-trans-I 1-ciseicosatetraenoic acid) stimulates secretion from leukocytes, activates protein kinase C, and induces vasodilation in the hamster cheek pouch and rat kidney. Both of LXA 4 and LXB 4 (5S, 14R, 15S-trihydroxy-6,10,I 2-trans-8-cis-eicosatetraenoic acid) inhibit cytotoxicity induced by natural killer cells. In addition, A 4 displays contractile activity in the guinea pig lung strip [3]. Here we summarize our present information about the mode of action of LXA 4 in this spasmogenic assay. The data indicate that LXA 4 in this model for pulmonary smooth muscle reactivity activates a site which is closely similar to, or even identical with, the receptor(s) for cysteinyl-containing leukotrienes (cysLTs, i.e. LTC4, LTD 4 and LTE4).

The Total Synthesis of the Lipoxins and Related Compounds

In 1984, a new class of arachidonic acid derived products was discovered by Serhan et al.1–3 While studying lipoxygenase pathway interactions, these researchers isolated two major constituents from human leukocytes. These substances displayed interesting biological properties including the ability to suppress the cytotoxic activity of natural killer cells.lb,lc,3–7