CGRP 27-37 analogues with high affinity to the CGRP1 receptor show antagonistic properties in a rat blood flow assay

CGRP Y-0-28-37 is known as a selective CGRP(1) receptor antagonist. We succeeded in optimising the CGRP(1) receptor affinity of this fragment by multiple amino acid replacement. The analogues [P-34, F-35]CGRP 27-37 and [D-31, p(34), F-35]CGRP 27-37 exhibit a 100-fold increased affinity compared to the unmodified segment. Receptor binding studies were performed with human neuroblastoma cells SK-N-MC, which selectively express the hCGRP(1) receptor. Blood flow, which is increased by exogenous CGRP, was measured in the right femoral artery. Preincubation of the rats with [P-34, F-35]CGRP 27-37 and [D-31, p(34), F-35]CGRP 27-37 led to a significant decrease in CGRP induced increase in vascular conductance indicating the antagonistic properties of these compounds. Interestingly, an exchange of the amino acid Asn(31) to Asp(31) in [P-34, F-35]CCRP 27-37 shortened the period of the antagonistic effect significantly, suggestive of a different rate of metabolism for the two ligands. Secondary structure investigations obtained by circular dichroism measurements revealed that an increase in ordered structure correlates with high binding affinity. (C) 1999 Elsevier Science B.V. All rights reserved.