SYNTHESIS, STRUCTURE, AND ANTAGONISTIC PROPERTIES OF DES-ASN(29)[D-TRP(28,32)]NPY(27-36)

We have previously reported that [D-Trp(32)]NPY and its centrally trun cated analogues such as des-AA(7-24)[D-Trp(5,32),Aoc(6)] NPY can compe titively antagonize NPY effects on rat hypothalamus and Y-1 (SK-N-MC A ND HEL) cells, respectively. In continuation of this work, we performe d structure-activity studies with C-terminal decapeptide sequence keep ing D-Trp at position 32 to develop lower molecular weight Y-1-selecti ve antagonists. This study led to the development of des-Asn(29)[D-Trp (28,32)]NPY(27-36), which bound to both Y-1 (SK-N-MC, K-i greater than or equal to 10 mu M) and Y-2 (SK-N-BE2, K-i = 1.01 +/- 0.03 mu M) rec eptors. This peptide did not exhibit any agonist activity at Y-1 recep tors, and exhibited comparable potencies in antagonizing the effects o f NPY on the synthesis of cAMP and mobilization of [Ca2+](i) in HEL ce lls. However, in SK-N-MC cells, it was more potent in antagonizing the mobilzation of [Ca2+](i) than inhibition of cAMP synthesis. Substitut ion of Nva for Gln(34) to increase the hydrophobicity without altering the carbon skeleton substantially increased Y-1 affinity (K-i = 0.33 +/- 0.15 mu M) and imparted Y-1 selectivity (K-i for Y-2 affinity = 3. 16 +/- 0.50). Moreover, this peptide exhibited good antagonistic poten cy in HEL cells. 2D NMR studies of des-Asn(29)[D-Trp(28,32)]NPY(27-36) revealed the existence of a fairly stable loop-like structure between residues 27 and 32 and a less stable one between residues 32 and 36. The increased Y-1 affinity of des-Asn(29)[D-Trp(28,32),Nva(34)]NPY(27- 36) may be due to the stabilization of the 32-36 loop by Nva(34). It a ppears therefore that stabilization of the loop structures in these pe ptides should result in the development of more potent Y-1 receptor an tagonists. Our investigations also suggest that HEL cells express a ho mogeneous population of NPY Y-1 receptors whereas SK-N-MC cells expres s high- and low-affinity Y-1 receptors coupled to Ca2+ and cAMP, respe ctively. Copyright (C) 1996 Elsevier Science Inc.