Allosteric lipoxygenase effectors as potential drugs

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Arachidonic acid lipoxygenases (ALOXs) and their metabolites (eicosanoids and related substances) have been implicated in cell maturation and differentiation, but also in the pathogenesis of inflammatory, hyperproliferative and neurological diseases. Over the past twenty years scientists have developed drugs preventing the biosynthesis of eicosanoids or inhibiting the interaction of these mediators with the corresponding receptors. Today anti-inflammatory COX2 inhibitors (COXIBs) are among the most selling drugs worldwide and inhibitors of the ALOX5 pathway as well as leukotriene receptor (cysLTR1) antagonists have been developed. Some of them (Zileuton, Montelukast, Zafirlukast) are currently available for prescription as anti-asthmatic drugs. Completion of the human genome project indicated the existence of six functional ALOX genes (ALOX5, ALOX15, ALOX15B, ALOX12, ALOX12B, ALOXE3), which encode for six different ALOX-isozymes. For five of them including ALOX15 and ALOX5 knockout mice are available and experiments with these animals indicated distinct biological functions for the different ALOX isoforms. For ALOX15 pro- and anti-inflammatory properties (dual role of ALOX15) have been reported and detailed kinetic investigations recently suggested that human ALOX15 exhibits allosteric properties. This finding was rather surprising since the crystal structures of mammalian ALOX15 orthologs have indicated that the enzymes consist of a single polypeptide chain and there was no evidence for a conserved allosteric ligand binding site. In aqueous solutions ALOX15 forms catalytically active dimers consisting of a catalytic monomer, which binds the substrate fatty acid at the active site, and a regulatory monomer serving as binding site for allosteric ligands. The allosteric character of the ALOX15 reaction offers the possibility for the development of isoform-specific ALOX15 effectors (inhibitors and activators), which are currently not available.When we explored the biological activities of several substituted indolylphenylaminosulfocarbamates we found them to selectively prevent ALOX15 dependent linoleic acid oxygenation. In contrast, oxidation of arachidonic acid was not inhibited. Detailed kinetic measurements, analysis of the reaction products and preliminary structural modelling on the basis of the ALOX15 X-ray coordinates suggested that these substrate specific effectors bind at the substrate-binding pocket of the allosteric monomer. This binding induces a conformational change of the second catalytic monomer, which inhibits oxygenation of linoleic acid but not of arachidonic acid. Together these data indicate the possibility for the development of substrate-specific allosteric ALOX15 effectors. Considering the dual functionality of ALOX15 in inflammation the search for allosteric ALOX15 effectors (inhibitors and activators) may be more promising than conventional inhibitor screening strategies.