β-Pro⁷Ang III is a novel highly selective angiotensin II type 2 receptor (AT₂R) agonist, which acts as a vasodepressor agent via the AT₂R in conscious spontaneously hypertensive rats

We have previously shown that individual β-amino acid substitution in angiotensin (Ang) II reduced Ang II type 1 receptor (AT₁R) but not Ang II type 2 receptor (AT₂R)-binding and that the heptapeptide Ang III exhibited greater AT₂R:AT₁R selectivity than Ang II. Therefore, we hypothesized that β-amino-acid-substituted Ang III peptide analogues would yield highly selective AT₂R ligands, which we have tested in binding and functional vascular assays. In competition binding experiments using either AT₁R- or AT₂R-transfected human embryonic kidney (HEK)-293 cells, novel β-substituted Ang III analogues lacked appreciable AT₁R affinity, whereas most compounds could fully displace ¹²⁵I-Sar¹Ile⁸ Ang II from AT₂R. The rank order of affinity at AT₂R was CGP42112 > Ang III > β-Pro⁷ Ang III=Ang II > β-Tyr⁴ Ang III ≥ PD123319 >> β-Phe⁸ Ang III >> β Arg² Ang III=β-Val³ Ang III >> β-Ile⁵ Ang III. The novel analogue β-Pro⁷ Ang III was the most selective AT₂R ligand tested, which was >20 000-fold more selective for AT₂R than AT₁R. IC₅₀ values at AT₂R from binding studies correlated with maximum vasorelaxation in mouse aortic rings. Given that β-Pro⁷ Ang III was an AT₂R agonist, we compared β-Pro⁷ Ang III and native Ang III for their ability to reduce blood pressure in separate groups of conscious spontaneously hypertensive rats. Whereas Ang III alone increased mean arterial pressure (MAP), β-Pro⁷ Ang III had no effect. During low-level AT₁R blockade, both Ang III and β-Pro⁷ Ang III, but not Ang II, lowered MAP (by ∼30 mmHg) at equimolar infusions (150 pmol/kg/min for 4 h) and these depressor effects were abolished by the co-administration of the AT₂R antagonist PD123319. Thus, β-Pro⁷ Ang III has remarkable AT₂R selectivity determined in binding and functional studies and will be a valuable research tool for insight into AT₂R function and for future drug development.