5-Oxo-ETE is the most powerful eosinophil chemoattractant among lipid mediators. Eosinophil infiltration into the lungs of asthmatics may be responsible for the late phase of inflammatory asthma. We have designed and synthesized a 5-oxo-ETE receptor antagonist, the purpose of which is to prevent eosinophil migration to the lung during an asthma attack and thereby reduce asthma symptoms.

Glutamate is an excitatory neurotransmitter that controls numerous pathways in the brain. Neuroscientists make use of photoremovable protecting groups, also known as cages, to release glutamate with precise spatial and temporal control. Various cage designs have been developed and among the most effective has been the nitroindolinyl caging of glutamate. We, hereby, report an improved synthesis of one of the current leading molecules of caged glutamate, 4-carboxymethoxy-5,7-dinitroindolinyl glutamate (CDNI-Glu), which possesses efficiencies with the highest reported quantum yield of at least 0.5. We present the shortest route, to date, for the synthesis of CDNI-Glu in 4 steps, with a total reaction time of 40 h and an overall yield of 20%. We also caged glutamate at the other two functional groups, thereby, introducing two new cage designs: α-CDNI-Glu and N-CDNI-Glu. We included a study of their photocleavage properties using UV-vis, NMR, as well as a physiology experiment of a two-photon uncaging of CDNI-Glu in acute hippocampal brain slices. The newly introduced cage designs may have the potential to minimize the interference that CDNI-Glu has with the GABA receptor. We are broadly disseminating this to enable neuroscientists to use these photoactivatable tools.

a b s t r a c tNitrated indolinyl photoprotecting groups are crucial tools extensively used in the study of neuronal signal transduction. Mononitrated photolabile protecting groups have been used effectively, however, recent advances in the introduction of a second nitro group have shown improvement in the photo efficiency of neurotransmitter (agonist) release, albeit, to varying extents, depending on the assessment methods employed. An unambiguous method is discussed based on Nuclear Magnetic Resonance (NMR), which is shown to be an effective technique in the relative overall rate comparison amongst varying nitrated protecting groups. Mononitrated and dinitrated photolabile protecting groups such as CDNIGlu and MNI-Glu are used as an example to assess the relative value of adding a second nitro group in photoactive cage designs. Using this technique, it was shown that the second nitro group in CDNI systems enhances the overall relative rate of photocleavage by a factor of 5.8. This reported method can also be used to unambiguously determine relative rate of agonist photorelease.

5-Oxo-6,8,11,14-eicosatetraenoic acid (5-oxo-ETE) is formed by the oxidation of 5-hydroxy-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-HETE), which is a major metabolite of enzymatic oxidation of arachidonic acid (AA). 5-Oxo-ETE is the most potent lipid chemoattractant for human eosinophils. Its actions are mediated by the selective OXE receptor, which is therefore an attractive target in eosinophilic diseases such as allergic rhinitis and asthma. Recently, we have reported two excellent OXE receptor antagonists that have IC 50 values at low nanomolar concentrations. Each of these antagonists has a chiral center, and the isolation of the individual enantiomers by chiral high-performance liquid chromatography (HPLC) revealed that in each case one enantiomer is over 300 times more potent than the other. To unambiguously assign the stereochemistry of these enantiomers and to provide access to larger amounts of the active compounds for biological testing, we report here their total synthesis. KEYWORDS: 5-Oxo-ETE, eosinophil, OXE receptor, antagonist, enantiomeric, optical purity, chiral auxiliary A sthma is a complex disease, which is essentially characterized by a bronchoconstriction. In most cases it is composed of an early phase (or acute) and a late-phase (or inflammatory) asthma. One of the hallmarks of the disease is the accumulation of eosinophils in the early phase, which is then responsible for the late-phase. 5-Oxo-ETE is a potent eosinophil chemotactic factor 1,2 and responsible for the accumulation of eosinophils in the lungs 3 (Figure 1).It is formed from arachidonic acid (AA) by the 5-lipoxygenase (5-LO) pathway, following oxidation of 5-hydroxy-6E,8Z,11Z,14Z-eicosatetraenoic acid (5-HETE) by 5-hydroxyeicosanoid dehydrogenase in the presence of NADP + (Figure 2). 4,5 After the first total synthesis of 5-oxo-ETE, 6 we have identified and investigated the activities of its metabolites on the 5-oxo-ETE receptor (OXE-R). 7 We found that a metabolite of 5-oxo-ETE formed by platelets, 5-oxo-12(S)-HETE, inhibits 5-oxo-ETE-induced Ca 2+ mobilization, though not increasing intracellular Ca 2+ concentration by itself. 8 However, this compound is not suitable for the development of an OXE-R antagonist as it is not stable under physiological conditions. Studies on a variety of 5-oxo-ETE analogues clearly showed that the carboxyl, carbonyl (including the 5-oxo group), and the alkyl regions of the molecule are essential for biological activity. 9 We therefore sought to develop synthetic OXE-R antagonists by incorporating both 5-oxo-valeryl and hexyl groups onto an indole backbone. The selection of the indole system looked more promising. We synthesized other aromatic scaffolds such as naphthalene, benzofuran, and quinoline, which turned out to be inactive. These studies culminated in the identification of two potent OXE-R antagonists (Figure 3) with IC 50 values of less than 30 nM. 10,11 Both of these compounds (5 and 6) have an asymmetric center on their acyl side chain, and it seemed likely that their activiti...