Inhibition and time-dependent inactivation of human placental aromatase by 3-oxo-17β-carboxamido steroids

Abstract

Several N, N-dialkyl-3-oxo-4-aza-17β-carboxamido steroids were found to be competitive inhibitors versus androstenedione (AND) and time-dependent inactivators of aromatase activity from human term placenta! microsomes. Inhibition constants ( K is ) from dead-end inhibition analyses indicated interactions between these compounds and the enzyme over a 0.8–7 μM inhibitor concentration range. The affinity of these compounds for aromatase leading to the time-dependent loss of enzyme activity was several fold higher than that estimated by the steady-state kinetics, with rate constants of inactivation of 0.025–0.033 min −1. 3-Oxo-4-aza steroids lacking a 17β-carboxamide were found to be competitive inhibitors of AND for aromatase, but did not inactivate enzyme activity in a time-dependent manner. Steroids which did not contain a 4-aza substituent, but retained the 17β-carbamoyl functionality, were both inhibitors and inactivators of aromatase activity in the microsomes. The time-dependent loss of aromatase activity induced by these compounds was shown to require reducing equivalents as provided by NADPH. Hence, it is suggested that the inactivation of aromatase by compounds in this series is dependent on enzymatic activation in the presence of the N, N-dialkyl-17β-carbamoyl substituent.