Postmortem protein stability investigations of the human hepatic drug-metabolizing cytochrome P450 enzymes CYP1A2 and CYP3A4 using mass spectrometry

摘要

Variability in expression and activity of hepatic drug-metabolizing cytochrome P450 (CYP) enzymes can play a causal role in fatal intoxication cases and is thus of forensic interest. We investigated the feasibility of LC-MS/MS based quantification and in vitro enzyme activity measurements of two major drug-metabolizing enzymes CYP1A2 and CYP3A4 in postmortem human liver microsomes (HLM). In autopsy cases (postmortem interval 24-36 h) we found CYP1A2 and CYP3A4 protein levels similar to that measured in a non-decayed reference HLM pool, whereas CYP1A2 and CYP3A4 enzyme activities were absent or severely decreased. Stability studies showed that CYP1A2 and CYP3A4 protein abundances were relatively stable in tissue stored in vitro for up to seven days at 4 degrees C. When tissue was stored for more than one day at 21 degrees C variable and case-specific decay patterns were observed, and CYP abundances declined especially after 3-4 days storage. Investigations of 50 autopsy cases revealed mean CYP1A2 and CYP3A4 levels of 49 and 47 pmol per mg HLM protein and inter-individual variabilities similar to those reported in other studies. This study supports postmortem quantification of CYP proteins in autopsy hepatic tissue by mass spectrometry.