CGRP 27–37 analogues with high affinity to the CGRP 1 receptor show antagonistic properties in a rat blood flow assay

Abstract

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]CGRP 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.