Metabolic stability and effects of microsomal enzymes on the activity of the sonic hedgehog pathway inhibitor piperonyl butoxide.

Abstract

The pesticide synergist piperonyl butoxide (PBO) is a methylenedioxy compound used in many pesticide formulations. Previous studies identified PBO as an inhibitor of the Sonic hedgehog (Shh) signaling pathway and linked prenatal PBO exposure to adverse developmental outcomes. Mixed-function oxidases have been proposed to metabolize PBO, but the specific enzymes involved in its depletion have not been identified. Here we examined the metabolic stability of PBO in the presence of human liver microsomes and the involvement of the CYP-450 (CYPs) and FMO enzyme families on the in vitro depletion of PBO. We found that PBO is readily depleted by microsomal enzymes in the presence of NADPH. The CYP inhibitor SKF-525 A significantly decreased PBO depletion, while the FMO inhibitor methimazole did not. We then examined the depletion capacity of individual CYPs, focusing on isoforms with common human polymorphisms. CYP2C19, CYP2C9, and CYP3A4 exhibited the greatest PBO depletion capacity, while CYP1A2 and CYP2D6 demonstrated moderate capacity. Finally, the effect of microsomal activity on the antagonist activity of PBO against the Sonic hedgehog (Shh) pathway was assessed. Microsomal depletion reduced but did not eliminate the antagonistic activity of PBO on Shh pathway signaling activity. Collectively, these findings suggest a major role for mixed-function oxidases in PBO depletion and indicate the possible involvement of specific CYP isoforms.