Both willardiine and azawillardiine analogs (18-28) have been reported to be potent and selective agonists for either AMPA or kainate receptors. We report here the novel synthesis and pharmacological characterization of a range of willardiine (18-23) and 6-azawillardiine (24-28) analogs on cells individually expressing human homomeric hGluR1, hGluR2, hGluR4, or hGluR5 receptors. Reaction of the sodium salts of substituted uracils (7-12) or 6-azauracils (13-16) with (S)-3-[(tert-butoxycarbonyl)amino]oxetan-2-one (17) in dry DMF, subsequent deprotection in TFA, and purification by ion-exchange chromatography gave mainly the willardiine analog in which alkylation took place on N1 of the uracil ring. We have investigated the subtype selectivity of these compounds by examining their binding affinity for homomeric hGluR1, -2, -4, or -5 (and hGluR6 in the case of 5-iodowillardiine (22)). From this study we have demonstrated that 22 has high affinity for hGluR5 and, compared to kainate, displays excellent selectivity for this receptor over both the AMPA receptor subtypes and the homomeric kainate receptor, hGluR6. 5-Fluorowillardiine (19) has higher affinity than AMPA for both homomeric hGluR1 and hGluR2 and compared to AMPA displays greater selectivity for AMPA receptor subtypes over the kainate receptor, hGluR5. Some structural features required for optimal activity at homomeric AMPA or kainate receptor subtypes have also been identified. It would appear that quite large lipophilic substituents at the 5-position of the uracil ring not only are accommodated by hGluR5 receptors but also lead to enhanced affinity for these receptors. In contrast to this, for optimal binding affinity to hGluR1, -2, or -4, smaller, electron-withdrawing substituents are required. For optimal activity at hGluR4 receptors a 6-aza-substituted willardiine is favored. The subtype-selective compounds described here are likely to be useful tools to probe the distribution and the physiological roles of the various glutamate receptor subunits in the central nervous system.

Benzylimidazolines may represent a class of 5-HT1D ligands that has yet to be exploited. On the basis of a previous report that the 2-(substituted-benzyl)imidazoline alpha-adrenergic agonist oxymetazoline (8) binds with high affinity at calf brain 5-HT1D receptors, we explored the structure-affinity relationships of a series of related derivatives. Each of the aromatic substituents was removed and then reinstated in a systematic manner to determine the influence of the individual substituents on binding. It was found that all of the aromatic substituents of 8 act in concert to impart high affinity. However, although the 3-hydroxy group could be removed without significantly reducing affinity for h5-HT1D (i.e., human 5-HT1Dalpha) receptors, this modification reduced h5-HT1B (i.e., human 5-HT1Dbeta) receptor affinity by nearly 50-fold. The 2, 6-dimethyl groups also contribute to binding but seem to play a greater role for h5-HT1B binding than h5-HT1D binding. With the appropriate structural modifications, several compounds were identified that display 20- to >100-fold selectivity for h5-HT1D versus h5-HT1B receptors. Preliminary functional data suggest that these compounds behave as agonists. Given that 5-HT1D agonists are currently being explored for their antimigraine action and that activation of h5-HT1B receptors might be associated with cardiovascular side effects, h5-HT1D-selective agents may offer a new lead for the development of therapeutically efficacious agents.

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