Antagonistic Properties of Centrally Truncated Analogs of [d-Trp32]NPY

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Abstract
We have previously shown that [d-Trp32]NPY can competitively antagonize NPY-induced feeding in rats (Balasubramaniam et al. J. Med. Chem. 1994, 37, 811−815). This peptide, however, did not bind to SK-N-MC cells with Y-1 receptors. Since centrally truncated NPY analogs have been shown to bind Y-1 receptors, we synthesized similar analogs of [d-Trp32]NPY and investigated their Y-1 (SK-N-MC) and Y-2 (SK-N-BE2) receptor affinities and their properties in human erythroleukemia (HEL) cells. None of the analogs with d-Trp32 mobilized intracellular calcium, [Ca2+]i, in HEL cells. Although Des-AA6-24[Aoc6]NPY and the corresponding d-Trp32 analog exhibited no affinity to Y-1 receptors, Des- AA7-24[Aoc6,d-Trp32]NPY (6) exhibited weak binding. Replacing Pro5 in 6 with d-Ala to stabilize the central chain reversal, and hence the antiparallel alignment of the N- and C-terminal regions known to be important for Y-1 binding, resulted in an analog, Des-AA7-24[d-Ala5,Aoc6,d-Trp32]NPY (7), which exhibited moderate antagonist potency in attenuating NPY effects on cAMP and [Ca2+]i in SK-N-MC and HEL cells, respectively. This analog also shifted the dose−response curve of NPY on blood pressure in anesthetized rats. Deletion of only the 7−17 and/or the incorporation of N-Me-Ala5, a superior β-turn stabilizer, in 7 did not improve the Y-1 receptor affinity. Des-AA7-24[d-Ala5,Gly6,d-Trp32]NPY exhibited an affinity similar to that of 7, suggesting that a long spacer arm is not necessary for efficient Y-1 receptor interaction. Locking the antiparallel alignment via a 2/26 or 2/27 lactam bridge did not improve the binding. Finally, replacement of d-Ala5 in 7 with d-Trp dramatically increased both the binding and the antagonistic potencies. Modeling based on the avian pancreatic polypeptide X-ray structure suggested that analogs which have the N- and C-terminal regions in close proximity might exhibit good binding, and that the d-Trp32 substitution may induce a β-turn that could be important for exhibiting antagonism. A systematic investigation has resulted in the development of relatively potent Y-1 receptor antagonists. Further structure−activity studies with these compounds and those previously reported by us and other investigators should result in the development of long-acting and receptor selective antagonists.