Elsevier

Peptides

Volume 18, Issue 3, 1997, Pages 403-408
Peptides

The Long-Acting Vasoactive Intestinal Polypeptide Agonist RO 25-1553 Is Highly Selective of the VIP2 Receptor Subclass

https://doi.org/10.1016/S0196-9781(96)00322-1Get rights and content

Abstract

Gourlet, P., P. Vertongen, A. Vandermeers, M.-C. Vandermeers-Piret, J. Rathe, P. De Neef, M. Waelbroeck and P. Robberecht. The long-acting vasoactive intestinal polypeptide agonist RO 25-1553 is highly selective of the VIP2 receptor subclass. Peptides 18(3) 403–408, 1997.—RO 25-1553 is a synthetic VIP analogue that induced a long-lasting relaxation of tracheal and bronchial smooth muscles as well as a reduction of edema and eosinophilic mobilization during pulmonary anaphylaxis. In the present study, we tested in vitro the capacity of RO 25-1553 to occupy the different VIP/PACAP receptor subclasses and to stimulate adenylate cyclase activity. The cellular models tested expressed one single receptor subtype: Chinese hamster ovary (CHO) cells transfected with the rat recombinant PACAP I, rat VIP1, and human VIP2 receptors; SUP T1 cells expressing the human VIP2 and HCT 15 and LoVo cells expressing the human VIP1 receptor. RO 25-1553 was threefold more potent than VIP on the human VIP2 receptor, 100- and 600-fold less potent than VIP on the rat and human VIP1 receptors, respectively, and 10-fold less potent than VIP and 3000-fold less potent than PACAP on the PACAP I receptor. RO 25-1553 was a full agonist on the VIP2, the PACAP I, and the rat recombinant VIP1 receptor but a partial agonist only on the human VIP1 receptor. Thus, RO 25-1553 is a highly selective agonist ligand for the VIP2 receptor subclass.

Section snippets

Peptide Synthesis

VIP, PACAP-27, and PACAP-38 were synthesized by solid-phase methodology using the Fmoc strategy. RO 25-1553 was synthesized as described [19]using the Nα-Boc derivatives. All the peptides were purified on various HPLC and FPLC columns. The purity of the material was at least 95% as judged by capillary electrophoresis and analytical reverse-phase chromatography and the peptide conformity was assessed by mass spectrometry.

Cell Cultures

Chinese hamster ovary cells (CHO cells) expressing the recombinant rat VIP1

Characteristics of the Cell Lines

The characteristics of the CHO cell lines transfected with the DNA coding for the rat VIP1, human VIP2, and rat PACAP type I receptors were the following: [125I]VIP recognized 850 ± 50 and 100 ± 30 fmol of rat VIP1 receptor/mg protein on VIP1 r clone 3 and 16 cells, respectively [6]; 210 ± 40 fmol of human VIP2 receptor/mg protein on the VIP2 r clone 11 cells [27], and [125I]PACAP-27 detected 4.6 ± 0.3 pmol of rat PACAP I receptor/mg protein on PACAP I r clone P2-10 cells [7]. mRNA was prepared

Discussion

VIP/PACAP receptors are classified as PACAP type I receptors with a high affinity for PACAP and a low affinity for VIP, and PACAP type II receptors, with an equal high affinity for PACAP and VIP. The PACAP type II receptors were further subdivided into VIP1 and VIP2 receptors 5, 21. VIP1 receptors have been cloned in rat [13]and human [8]; VIP2 receptors have been cloned in rat [17], mouse [12], and human [27]. The VIP2 receptor corresponds to the “helodermin-preferring” receptor previously

Acknowledgements

Aided by Grant No. 3.4502.95 from the Fonds de la Recherche Scientifique Médicale and by an Action Concertée de Recherche from the Communauté Française de Belgique.

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