The Broader Context of Liquid Biopsy in Absorption, Distribution, Metabolism, and Elimination.

We read with interest the manuscript of Pridgeon and colleagues entitled “Liquid Biopsies or Therapeutic Drug Monitoring for CYP Activity Profile Determination.” Given the title of the manuscript it is notable that no experiments evaluating the performance of either liquid biopsy or therapeutic drug monitoring (TDM) approaches to determine cytochrome P450 (CYP) activity are described. Rather the manuscript describes a set of in vitro analyses that confirm a weak correlation between messenger RNA (mRNA) and protein expression for CYP within human liver tissue. By extrapolation, the authors infer but do not demonstrate poor performance for mRNAbased liquid biopsies. We acknowledge and agree that confirmation of prior findings in human liver tissue of the relatively weak correlation between CYP mRNA and protein expression does highlight the need for validation studies for mRNAbased liquid biopsies. However, Pridgeon and colleagues conflate the general concept of a liquid biopsy with one specific implementation using mRNA. While the results reported in the manuscript are pertinent to mRNA liquid biopsies, the findings cannot be appropriately interpreted without context highlighting the existence of alternative liquid biopsy approaches for assessment of CYP activity that are not based on measurement of mRNA. Notably, mRNA is only one type of cargo contained within extracellular vesicles that may serve as a marker of tissue CYP activity. Specifically the authors make no reference to the growing evidence demonstrating the presence and performance of extracellular vesicle– derived protein expression and activity with respect to characterizing variability in exposure to medicines cleared by CYP. 5 In doing so the authors underplay the current stateoftheart with respect to the potential application of liquid biopsy for the determination of in vivo CYP activity. Although not mentioned in the manuscript, CYP protein can be quantified in plasma extracellular vesicles. Additionally, the activity of CYP3A4 protein in plasma extracellular vesicles can be quantified using midazolam as a probe substrate. Both CYP3A4 protein and CYP3A4 activity in plasma extracellular vesicles have demonstrated strong correlation with in vivo CYP3A4 activity. Notably, the issue of poor mRNA correlation reported is not relevant for these types of liquid biopsies. Nonetheless, extensive validation studies are still required before such an approach could be considered for guiding clinical dosing. Finally, it is worth noting that while liquid biopsy, pharmacogenomics, and TDM are technologies to support precision dosing, they serve distinct complementary functions and do not need to be viewed as competing technologies. TDM is used to inform ontreatment dose adjustments, whereas liquid biopsy may complement pharmacogenomics to inform optimal initial dose selection and help to identify subgroups of patients for whom ontreatment monitoring is likely to be most important.