Major glucuronide metabolites of testosterone are primarily transported by MRP2 and MRP3 in human liver, intestine and kidney

Abstract

Testosterone glucuronide (TG), androsterone glucuronide (AG), etiocholanolone glucuronide (EtioG) and dihydrotestosterone glucuronide (DHTG) are the major metabolites of testosterone (T), which are excreted in urine and bile. Glucuronides can be deconjugated to active androgen in gut lumen after biliary excretion, which in turn can affect physiological levels of androgens. The goal of this study was to quantitatively characterize the mechanisms by which TG, AG, EtioG and DHTG are eliminated from liver, intestine, and kidney utilizing relative expression factor (REF) approach. Using vesicular transport assay with recombinant human MRP2, MRP3, MRP4, MDR1 and BCRP, we first identified that TG, AG, EtioG, and DHTG were primarily substrates of MRP2 and MRP3, although lower levels of transport were also observed with MDR1 and BCRP vesicles. The transport kinetic analyses revealed higher intrinsic clearances of TG by MRP2 and MRP3 as compared to that of DHTG, AG, and EtioG. MRP3 exhibited higher affinity for the transport of the studied glucuronides than MRP2. We next quantified the protein abundances of these efflux transporters in vesicles and compared the same with pooled total membrane fractions isolated from human tissues by quantitative LC-MS/MS proteomics. The fractional contribution of individual transporters (f t ) was estimated by proteomics-based physiological scaling factors, i.e., transporter abundance in whole tissue versus vesicles, and corrected for inside-out vesicles (determined by 5'-nucleotidase assay). The glucuronide metabolites of inactive androgens, AG and EtioG were preferentially transported by MRP3, whereas the glucuronides of active androgens, TG and DHTG were mainly transported by MRP2 in liver. Efflux by bile canalicular transport may indicate the potential role of enterohepatic recirculation in regulating the circulating active androgens after deconjugation in the gut. In intestine, MRP3 possibly contributes most to the efflux of these glucuronides. In kidney, all studied glucuronides seemed to be preferentially effluxed by MRP2 and MDR1 (for EtioG). These REF based analysis need to be confirmed with in vivo findings. Overall, characterization of the efflux mechanisms of T glucuronide metabolites is important for predicting the androgen disposition and interindividual variability, including drugandrogen interaction in humans. The mechanistic data can be extrapolated to other androgen

Human efflux transport of testosterone, epitestosterone and other androgen glucuronides

“…While our manuscript was under preparation, Li and colleagues (2019) published transport data on androsterone-G, dihydrotestosterone glucuronide, etiocholanolone-G and testosterone-G [44]. Our results are in line with their observations that MRP2 is a low affinity and MRP3 a high affinity transporter for androgen glucuronides, while MRP4 and BCRP do not transport these conjugates.…”

Quantitative Proteomics of Clinically Relevant Drug-Metabolizing Enzymes and Drug Transporters and Their Intercorrelations in the Human Small Intestine

“…synergistically, leading to an efficient first-pass drug extraction in the intestine (Cummins et al, 2002). There is also evidence associating the transmembrane transport of hydrophilic glucuronide metabolites formed by UGTs via MRP2 (Li et al, 2019); therefore, these correlations have some physiologic basis. A strong correlation observed between the P-gp and OST-a abundances in the gut wall (R 2 5 0.77) is only expected if they are regulated by the same nuclear receptors and/or if they share a common regulatory pathway (Tanigawara et al, 1999;Kast et al, 2002;Luo et al, 2002;Trubetskoy et al, 2005;Urquhart et al, 2007).…”

Unraveling the functional role of the orphan solute carrier, SLC22A24 in the transport of steroid conjugates through metabolomic and genome-wide association studies

“…The presence of this motif supports the direction of the observed associations and the idea that the transporter functions in the reabsorption of steroid conjugates in the kidney. Although it has been shown that the ABC transporters in the proximal tubule, MRP2 and MRP3 function in the efflux of a steroid conjugates into the urine and plasma, respectively[76], to our knowledge, there have been no reports characterizing renal reabsorptive SLC transporters for steroid glucuronide conjugates. Glucuronide conjugates of steroids normally undergo net secretion in the proximal tubule, presumably by MRP2 and the organic anion transporters, OAT1 and OAT3 [76].…”

“…Although it has been shown that the ABC transporters in the proximal tubule, MRP2 and MRP3 function in the efflux of a steroid conjugates into the urine and plasma, respectively[76], to our knowledge, there have been no reports characterizing renal reabsorptive SLC transporters for steroid glucuronide conjugates. Glucuronide conjugates of steroids normally undergo net secretion in the proximal tubule, presumably by MRP2 and the organic anion transporters, OAT1 and OAT3 [76]. Though our data suggest that SLC22A24 functions in the reabsorption of steroid metabolites in the kidney, future studies are needed to confirm its role.…”