Plasma extracellular nanovesicle (exosome)-derived biomarkers for drug metabolism pathways: a novel approach to characterize variability in drug exposure.

AIMS: Demonstrate the presence of cytochrome P450 (CYP) and UDP-glucuronosyltransferase (UGT) proteins and mRNAs in isolated human plasma exosomes and evaluate the capacity for exosome-derived biomarkers to characterize variability in CYP3A4 activity. METHODS: The presence of CYP and UGT protein and mRNA in exosomes isolated from human plasma and HepaRG cell culture medium was determined by mass spectrometry and reverse transcription-polymerase chain reaction, respectively. The concordance between exosome-derived CYP3A4 biomarkers and midazolam apparent oral clearance (CL/F) was evaluated in a small proof-of-concept study involving six genotyped (CYP3A4 *1/*1 and CYP3A5 *3/*3) Caucasian males. RESULTS: Exosomes isolated from human plasma contained peptides and mRNA originating from CYP 1A2, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, 2 J2, 3A4 and 3A5, UGT 1A1, 1A3, 1A4, 1A6, 1A9, 2B4, 2B7, 2B10 and 2B15, and NADPH-cytochrome P450 reductase. Mean (95% confidence interval) exosome-derived CYP3A4 protein expression pre- and post-rifampicin dosing was 0.24 (0.2-0.28) and 0.42 (0.21-0.65) ng ml-1 exosome concentrate. Mean (95% confidence interval) exosome CYP3A4 mRNA expression pre- and post-rifampicin dosing was 6.0 (1.1-32.7) and 48.3 (11.3-104) × 10-11 2-ΔΔCt , respectively. R2 values for correlations of exosome-derived CYP3A4 protein expression, CYP3A4 mRNA expression, and ex vivo CYP3A4 activity with midazolam CL/F were 0.905, 0.787 and 0.832, respectively. CONCLUSIONS: Consistent strong concordance was observed between exosome-derived CYP3A4 biomarkers and midazolam CL/F. The significance of these results is that CYP3A4 is the drug-metabolizing enzyme of greatest clinical importance and variability in CYP3A4 activity is poorly described by existing precision dosing strategies.