Identification of a crucial amino acid in the helix position 6.51 of human tachykinin neurokinin _1 and _3 receptors contributing to the insurmountable mode of antagonism by dual NK 1/NK 3 antagonists

摘要

The neurokinins are neuropeptides that elicit their effect through three GPCRs called NK _1, NK _2, and NK _3. Compounds 5 and 6 are dual hNK _1 (K _i of 0.7 and 0.3 nM) and hNK _3 (K _i of 2.9 and 1.7 nM) antagonists. Both compounds exhibit an insurmountable mode of antagonism at hNK 1, whereas at hNK 3, they differ in that 5 is an insurmountable but 6 a surmountable antagonist. Using homology modeling and site-directed mutagenesis, hNK 1-Phe264 and hNK 3-Tyr315 were found to be the molecular determinants of hNK _1 and hNK _3 antagonism by 5 and 6. In [3H]IP studies, the mutation hNK 1-F264Y converted the mode of action of 5 from insurmountable to partial insurmountable antagonism while it had no effect on that of 6. Conversely, the mutation hNK _3-Y315F enhanced the insurmountable behavior of 5 and converted 6's surmountable to an insurmountable antagonism. This finding was further confirmed by characterizing additional derivatives of 5 and 6, most notably with a hybrid structure.