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Pseudopterosin E (PSE), a C-10 linked fucose glycoside and pseudopterosin A (PSA), a C-9 xylose glycoside isolated from the marine gorgonian Pseudopterogorgia elisabethae were both effective in reducing PMA-induced mouse ear edema when administered topically (ED50 (microg/ear) PSE(38), PSA(8)) or systemically (ED50 (mg/kg, i.p.) PSE (14), PSA (32)). Both compounds exhibited in vivo analgesic activity in phenyl-p-benzoquinone-induced writhing (ED50 (mg/kg, i.p.) PSE(14), PSA(4). PSE inhibited zymosan-induced writhing (ED50 = 6 mg/kg, i.p.), with a concomitant dose-dependent inhibition of peritoneal exudate 6-keto-prostaglandin F1alpha (ED50 = 24 mg/kg) and leukotriene C4 (ED50 = 24 mg/kg). In vitro, the pseudopterosins were inactive as inhibitors of phospholipase A2, cyclooxygenase, cytokine release, or as regulators of adhesion molecule expression. PSA inhibited prostaglandin E2 and leukotriene C4 production in zymosan-stimulated murine peritoneal macrophages (IC50 = 4 microM and 1 microM, respectively); however, PSE was much less effective. These data suggest that the pseudopterosins may mediate their anti-inflammatory effects by inhibiting eicosanoid release from inflammatory cells in a concentration and dose-dependent manner.

Fuscoside (FSD) is a potent and long-lasting anti-inflammatory drug that selectively inhibits leukotriene production in murine models of inflammation. In the present study, the effects of FSD on the lipoxygenase pathways in human polymorphonuclear leukocytes are explored in order to better understand the mechanism of action of this novel drug. In adherent and suspended polymorphonuclear leukocytes, FSD irreversibly inhibits leukotriene B4 (LTB4) synthesis (IC50 = 10 microM) and the release of 14C-labeled LTB4 from neutrophils prelabeled with [14C]arachidonic acid. Unlike the reversible 5-lipoxygenase inhibitor L-651,896, FSD has no observable effect on LTB4 biosynthesis in whole blood, but does express activity as blood is successively diluted. In 10,000 x g supernatants of human platelets and polymorphonuclear leukocytes, FSD does not inhibit platelet 12-lipoxygenase, but is extremely effective in inhibiting the metabolism of arachidonic acid and 5-hydroperoxyeicosatetraenoic acid to LTB4 via neutrophil 5-lipoxygenase. FSD has no effect on the conversion of leukotriene A4 to LTB4 in this system. Interestingly, concurrent with FSD inhibition of leukotriene synthesis is a concentration-dependent increase in 5-hydroxyeicosatetraenoic acid, suggesting that FSD may selectively inhibit the leukotriene A4 synthase activity associated with human 5-lipoxygenase. FSD is therefore representative of a new class of nonantioxidant 5-lipoxygenase inhibitors that may be effective local therapeutic agents in the management of diseases such as psoriasis, arthritis and inflammatory bowel and lung diseases.

The biological and biochemical pharmacology of fuscoside, a novel anti-inflammatory marine natural product isolated from the Caribbean gorgonian Eunicea fusca, has recently been characterized using murine (part I) and human (part II) models of inflammation. Topically applied fuscoside (FSD) effectively inhibits phorbol myristate acetate (PMA)-induced edema in mouse ears at levels comparable with indomethacin over a 3.3-hr exposure period, and is significantly more efficacious than indomethacin over 24 hr in the PMA model. Histological preparations and quantification of the neutrophil-specific marker, myeloperoxidase, demonstrate that FSD inhibits neutrophil infiltration into PMA-induced regions of edema and inflammation. In systemic studies, where FSD is injected i.p. before the topical application of PMA, negligible effects on ear inflammation are observed. FSD does not inhibit bee venom or human synovial fluid phospholipase A2 up to concentrations of 500 microM. In calcium ionophore-activated cultures of mouse peritoneal macrophages, FSD selectively and irreversibly inhibits leukotriene C4 biosynthesis (IC50 = 8 microM), yet has negligible effects on prostaglandin E2 production. FSD is also without effect on the conversion of arachidonic acid to prostaglandin E2 by ram seminal vesicle cyclooxygenase. Chromatographic and spectroscopic studies suggest that FSD is not metabolized, and that drug uptake/binding by macrophages is time dependent, saturable and independent of active transport mechanisms. These studies represent the first report of an anti-inflammatory marine natural product that selectively inhibits leukotriene biosynthesis.

Brianolide (1), a new antiinflammatory diterpenoid of the briarein class, possessing a beta substituent at C-12 (R), has been isolated from the Okinawan gorgonian Briareum sp. Its structure has been established from spectral data in conjunction with a single crystal X-ray analysis.

The marine natural product manoalide (MLD), a potent inhibitor of phospholipases, completely inactivates bee venom phospholipase A2 (PLA2) by an irreversible mechanism. It has been proposed [K. B. Glaser and R. S. Jacobs, Biochem. Pharmac. 36, 2079 (1987)] that the reaction of MLD with PLA2 may involve the selective reactivity of MLD to a peptide sequence, possibly a Lys-X-X-Lys peptide. Localization of the MLD binding site on bee venom PLA2 demonstrated that upon MLD modification of bee venom PLA2 the only change in amino acid content was an apparent loss of Lys, corresponding to approximately three of the eleven Lys residues present. Selective chemical modification of Lys residues with [14C]maleic anhydride demonstrated that all eleven Lys residues on bee venom PLA2 were accessible to this reagent (11.6 mol maleyl group incorporated/mol of PLA2). Pretreatment of PLA2 with MLD (less than 0.7% residual activity) resulted in a molar ratio of 8.7, also consistent with the loss of three Lys residues upon modification by MLD. Reverse phase high performance liquid chromatography (RP-HPLC) of the cyanogen bromide (CNBr) digestion product of MLD-treated PLA2 produced three peaks (A280). The second peak showed the most intense absorbance at 434 nm. This material corresponded to residues 81-128, as determined by gas-phase microsequence analysis. Sequencing failure was observed at Lys-88 in the MLD-treated fragment. The control carboxymethylated-PLA2 fragment corresponding to residues 81-128 sequenced beyond Lys-88 without significant change in the expected yield. These data suggest that Lys-88 may correspond to one of the three MLD-modified Lys residues. The minor absorbance at 434 nm of the CNBr fragments containing residues 42-80 and 1-36 as compared to the fragment of residues 81-128 suggests that the major MLD binding fragment residues in residues 81-128.