N- and C-terminal structure-activity study of angiotensin II on the angiotensin AT2 receptor

The predominant angiotensin II receptor expressed in the human myometrium is the angiotensin AT2 receptor. This preparation was used for a structure-activity relationship study on angiotensin II analogues modified in positions 1 and 8. The angiotensin AT2 receptor present on human myometrium membranes displayed a high affinity (pKd = 9.18) and was relatively abundant (53-253 fmol/mg of protein). The pharmacological profile was typical of an angiotensin AT2 receptor with the following order of affinities: (angiotensin III > or = angiotensin II > angiotensin I > PD123319 > angiotensin-(1-7) > angiotensin-(1-6) approximately angiotensin IV >> Losartan). Modifications of the N-terminal side chain and of the primary amine of angiotensin II were evaluated. Neutralisation of the methylcarboxylate (Asp) to a methylcarboxamide (Asn) or to a hydroxymethyl (Ser) or substitution for a methylsulfonate group (cysteic acid) improved the affinity. Extension from methylcarboxylate (Asp) to ethylcarboxylate (Glu) did not affect the affinity. Introduction of larger side chains such as the bulky p-benzoylphenylalanine (p-Bpa) or the positively charged Lys did not substantially affect the affinity. Complete removal of the side chain (angiotensin III), however, resulted in a significant affinity increase. Removal or acetylation of the primary amine of angiotensin II did not noticeably influence the affinity. Progressive alkylation of the primary amine significantly increased the affinity, betain structures being the most potent. It appears that quite important differences exist between the angiotensin AT1 and AT2 receptors concerning their pharmacological profile towards analogues of angiotensin II modified in position 1. On position 8 of angiotensin II, a structure-activity relationship on the angiotensin AT2 receptor was quite similar to that observed with angiotensin AT1 receptor. Bulky, hydrophobic aromatic residues displayed affinities similar to or even better than [Sarcosine1]angiotensin II. Aliphatic residues, especially those of reduced size, caused a significant decrease in affinity especially [Sarcosine1, Gly8]angiotensin II who showed a 30-fold decrease. Introduction of a positive charge (Lys) at position 8 reduced the affinity even further. Stereoisomers in position 8 (L-->D configuration) also induced lower affinities. The angiotensin AT2 receptor display a structure-activity relationship similar to that observed on the AT1 receptor for the C-terminal position of the peptide hormone. Position 1 structure-activity relationships are however fundamentally different between the angiotensin AT1 and AT2 receptor.