Receptor binding affinity and biological activity of C-terminal elongated forms of endothelin-1

Endothelin-1 (21 amino acids; ET-21) is considered to be derived from a precursor, proendothelin (38 amino acids; ET-38). In order to make the physiological significance of this conversion clear, we synthesized various C-terminal elongated derivatives of ET-21, such as ET-22, ET-23, ET-25, ET-31, ET-36 and ET-38 (each number implies the number of amino acid residues), and measured their receptor binding affinities and biological activities. When inhibition of [ 125I]ET-21 binding to cultured rat smooth muscle cells (A10 cells) was measured, ET-21 inhibited with the highest affinity (IC 50 = 1.6 × 10 −10 M) and the affinity of ET-38 was 30-fold less than that of ET-21. The binding affinities of the C-terminal elongated peptides were reduced with increasing number of amino acid residues, except for ET-22 whose affinity was lower than those of other peptides (IC 50 = 1.6 × 10 −8 M). When contractions of rat aortic segments induced by these peptides were measured, ET-21 was the most potent (EC 50 = 2.8 × 10 −10 M). All C-terminal elongated peptides, including ET-38, were more than 100-fold less active. It is noteworthy that ET-22 was the least potent peptide (EC 50 = 1.2 × 10 −7 M). When bolus doses of C-terminal elongated peptides were administered to chemically denervated rats, the time-dependent change in blood pressure induced by each peptide was different from that induced by ET-21. Although ET-21 elicited a three phase depressor/pressor blood pressure response (an initial rapid hypotension, then a rapid transient hypertension followed by a slowly developing long-lasting hypertensive effect), the C-terminal elongated peptides, including ET-38, did not cause the initial transient hypotensive response. Very interestingly, the ability of the peptides to induce the rapid phase of hypertension in vivo does not seem to be correlated with the affinity of each peptide for the smooth muscle cell receptor, since the peptides with lower affinities for the smooth muscle receptor, such as ET-22, ET-23 and ET-25, showed more potent hypertensive effects. On the other hand, the slow and long-lasting hypertensive effect is likely to be related to the affinity of the compounds. The maximal hypertensive effects of cumulatively administered ET-21 derivatives were similar to those of ET-21. These results suggest that ET-21 is the most potent vasoconstrictor among the peptides and that the conversion from ET-38 to ET-21 may be important as an activation process.