Promiscuity and tissue-specific fractional contribution of UGT2B17 to drug and steroid metabolism
Creators: name: Ahire, Deepak
Publisher: name: Washington State University
Year: 2024
Subjects: Liquid chromatography-mass spectrometry
Non-CYP enzymes
Physiologically-based pharmacokinetics modeling
Quantitative Proteomics
UGT2B17
Uridine 5’-diphospho-glucuronosyltransferases

description: Oral drug administration is the preferred route of drug delivery. However, incomplete and variable bioavailability of drugs is often associated with unpredictable systemic drug exposure and efficacy. Such interindividual variability is associated with various factors such as age, genetics, epigenetics, disease conditions, sex, and drug-drug interactions (DDI) or natural product-drug interactions. Uridine 5'-diphospho-glucuronosyltransferase (UGT) 2B17 is one of the most highly polymorphic intestinal drug-metabolizing enzymes (DMEs) in its expression and activity. UGT2B17 variability contributes to inconsistent pharmacokinetics (PK) or efficacy of clinically important newer drugs such as verinurad, belzutifan, and asciminib. Because UGT2B17 is a minor isoform in the liver, the role of UGT2B17 in drug disposition is not well-studied for drugs developed before 2010 (e.g., diclofenac). Moreover, the structure-activity relationship of UGT2B17 substrates and inhibitors and the quantitative role of intestinal UGT2B17 in drug metabolism is unknown. This dissertation project focused on characterizing the promiscuity of UGT2B17 in drug and steroid metabolism and the contribution of this enzyme to the extensive and variable metabolism of drugs and steroids. Chapter 1 describes the significance, knowledge gaps, hypothesis, and specific aims of the work. Chapter 2 focuses on the development and validation of protein quantification methods for determining the abundance of DMEs and transporter proteins in human tissues. A microflow-based liquid chromatography-tandem mass spectrometry (µLC-MS/MS) method was developed for DMEs and transporter protein quantification using limited sample volume or protein concentration of liver tissues (1-100 mg), hepatocyte count (~4000 to 1 million cells), and microsomal protein concentration (0.01-1 mg/ml). Chapter 3 describes a novel universal conserved-plus-surrogate peptide (CPSP) approach for determining the accurate composition and total or cumulative abundance of homologous proteins in human tissues. UGT2B17 abundance data obtained using the CPSP approach was further used in Chapter 5 and Chapter 6 to calculate the tissue-specific fraction glucuronidated (fgluc) of diclofenac and vorinostat. To better explain the variability in vorinostat metabolism, ontogeny and variability in mitochondrial amidoxime-reducing component (mARC) enzymes (Chapter 4) were determined in human tissues. Chapter 5 characterized the promiscuity and quantitative contribution of UGT2B17 to drug and steroid metabolism. This study provided novel insight into the ligand binding interaction at the UGT2B17 active site and the tissue-specific fractional contribution of UGT2B17 to drug metabolism. Finally, the effects of UGT2B17 polymorphisms on diclofenac oral pharmacokinetics (PK) was predicted using a proteomics-informed physiologically-based pharmacokinetic (PBPK) model (Chapter 6).
description: In summary, this dissertation work revealed that UGT2B17 is a promiscuous enzyme and that the combination of highly variable and abundant intestinal UGT2B17 expression leads to the unpredictable PK of its substrates. Integration of the differential tissue abundance and