Abstract 169: Angiotensin-Converting Enzyme 2: Species-Specific Features and Pharmacologic Characterization by RAS Fingerprinting

Abstract

The carboxypeptidase Angiotensin Converting Enzyme 2 (ACE2) is a crucial player of the Renin-Angiotensin-System (RAS) as it balances the ratio between vasoconstrictive and fibrotic Angiotensin II and vasodilative and anti-fibrotic Angiotensin 1-7. This balance is affected by ACE2 either by direct conversion of Angiotensin II to Angiotensin 1-7, or by cleavage of Angiotensin I to Angiotensin 1-9, which is further converted to Ang 1-7 by Angiotensin Converting Enzyme (ACE) or Neutral Endopeptidase (NEP). An increased activity of ACE2 leads to a shift of the RAS towards an Angiotensin 1-7 dominated pattern, which was shown to be protective in various models of cardiovascular and fibrotic diseases. We performed a comparative pharmacologic characterization of recombinant murine and human ACE2 using natural peptide substrates for the evaluation of the substrate specific ACE2 activity in whole blood. Furthermore, we investigated the impact of ACE2 activators and inhibitors on murine and human ACE2 in whole blood using an LC-MS/MS based approach allowing the simultaneous quantification of 10 different angiotensin peptides (RAS-Fingerprinting). The investigation of inhibitors and activators of ACE2 using our experimental setting revealed the presence of species-specific enzymatic features, which was indicated by differences between the human and murine enzyme regarding the susceptibility to pharmacologic manipulation and the specificity for different endogenous peptide substrates. Our studies demonstrate that sequence diversity observed between recombinant human and murine ACE2 significantly affects substrate specificity and pharmacologic enzyme properties. The utilization of Angiotensin I by ACE2 generates Angiotensin 1-7 via Angiotensin 1-9 and represents a pathway of alternative RAS activation, which might have been systematically underestimated in previous studies. The strongly reduced affinity of murine ACE2 for Angiotensin I results in a lack of production of Angiotensin 1-7 via this pathway, which might be of particular importance during anti-hypertensive treatments with ACE inhibitors, where the formation of Angiotensin II is prevented while increased levels of Angiotensin I are observed in circulation.