The transport of monocarboxylates, such as lactate and pyruvate, is mediated by the SLC16A family of proton-linked membrane transport proteins known as monocarboxylate transporters (MCTs). Fourteen MCT-related genes have been identified in mammals and of these seven MCTs have been functionally characterized. Despite their sequence homology, only MCT1-4 have been demonstrated to be proton-dependent transporters of monocarboxylic acids. MCT6, MCT8 and MCT10 have been demonstrated to transport diuretics, thyroid hormones and aromatic amino acids, respectively. MCT1-4 vary in their regulation, tissue distribution and substrate/inhibitor specificity with MCT1 being the most extensively characterized isoform. Emerging evidence suggests that in addition to endogenous substrates, MCTs are involved in the transport of pharmaceutical agents, including γ-hydroxybuytrate (GHB), 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitors (statins), salicylic acid, and bumetanide. MCTs are expressed in a wide range of tissues including the liver, intestine, kidney and brain, and as such they have the potential to impact a number of processes contributing to the disposition of xenobiotic substrates. GHB has been extensively studied as a pharmaceutical substrate of MCTs; the renal clearance of GHB is dose-dependent with saturation of MCT-mediated reabsorption at high doses. Concomitant administration of GHB and l-lactate to rats results in an approximately two-fold increase in GHB renal clearance suggesting that inhibition of MCT1-mediated reabsorption of GHB may be an effective strategy for increasing renal and total GHB elimination in overdose situations. Further studies are required to more clearly define the role of MCTs on drug disposition and the potential for MCT-mediated detoxification strategies in GHB overdose.
butyrate; gamma-hydroxybutyrate; lactate; monocarboxylate transporters; SLC16A
The transport of monocarboxylates, such as lactate and pyruvate, is mediated by the SLC16A family of proton-linked membrane transport proteins known as monocarboxylate transporters (MCTs). Fourteen MCT-related genes have been identified in mammals and of these seven MCTs have been functionally characterized. Despite their sequence homology, only MCT1–4 have been demonstrated to be proton-dependent transporters of monocarboxylic acids. MCT6, MCT8 and MCT10 have been demonstrated to transport diuretics, thyroid hormones and aromatic amino acids, respectively. MCT1–4 vary in their regulation, tissue distribution and substrate/inhibitor specificity with MCT1 being the most extensively characterized isoform. Emerging evidence suggests that in addition to endogenous substrates, MCTs are involved in the transport of pharmaceutical agents, including γ-hydroxybuytrate (GHB), 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitors (statins), salicylic acid, and bumetanide. MCTs are expressed in a wide range of tissues including the liver, intestine, kidney and brain, and as such they have the potential to impact a number of processes contributing to the disposition of xenobiotic substrates. GHB has been extensively studied as a pharmaceutical substrate of MCTs; the renal clearance of GHB is dose-dependent with saturation of MCT-mediated reabsorption at high doses. Concomitant administration of GHB and l-lactate to rats results in an approximately two-fold increase in GHB renal clearance suggesting that inhibition of MCT1-mediated reabsorption of GHB may be an effective strategy for increasing renal and total GHB elimination in overdose situations. Further studies are required to more clearly define the role of MCTs on drug disposition and the potential for MCT-mediated detoxification strategies in GHB overdose.
butyrate; gamma-hydroxybutyrate; lactate; monocarboxylate transporters; SLC16A
The objective of the present study was to evaluate mechanistic pharmacokinetic models describing active renal secretion and reabsorption over a range of Michaelis–Menten parameter estimates and doses. Plasma concentration and urinary excretion profiles were simulated and renal clearance (CLr) was calculated for two pharmacokinetic models describing active renal reabsorption (R1/R2), two models describing active secretion (S1/S2), and a model containing both processes. A range of doses (1–1,000 mg/kg) was evaluated, and Vmax and Km parameter estimates were varied over a 100-fold range. Similar CLr values were predicted for reabsorption models (R1/R2) with variations in Vmax and Km. Tubular secretion models (S1/S2) yielded similar relationships between Michaelis–Menten parameter perturbations and CLr, but the predicted CLr values were threefold higher for model S1. For both reabsorption and secretion models, the greatest changes in CLr were observed with perturbations in Vmax, suggesting the need for an accurate estimate of this parameter. When intrinsic clearance was substituted for Michaelis–Menten parameters, it failed to predict similar CLr values even within the linear range. For models S1 and S2, renal secretion was predominant at low doses, whereas renal clearance was driven by fraction unbound in plasma at high doses. Simulations demonstrated the importance of Michaelis–Menten parameter estimates (especially Vmax) for determining CLr. Km estimates can easily be obtained directly from in vitro studies. However, additional scaling of in vitro Vmax estimates using in vitro/in vivo extrapolation methods are required to incorporate these parameters into pharmacokinetic models.
Electronic supplementary material
The online version of this article (doi:10.1208/s12248-012-9437-3) contains supplementary material, which is available to authorized users.
kidney transport parameters; models; reabsorption; renal clearance; secretion
The subcutaneous (SC) route is of growing interest for the administration of biotherapeutics. Key products on the biotherapeutic market such as insulins, but also several immunoglobulins or Fc-fusion proteins, are administered SC. Despite the importance of the SC route, the available knowledge about the processes involved in the SC absorption of biotherapeutics is limited. This review summarizes available information on the physiology of the SC tissue and on the pharmacokinetic processes after SC administration including “first pass catabolism” at the administration site as well as transport in the extracellular matrix of the SC tissue, followed by absorption into the blood circulation or the lymphatic system. Both monoclonal antibodies and other biotherapeutics are discussed. Determinants of absorption after SC administration are summarized including compound properties such as charge or molecular weight. Scale-up of animal data to humans is discussed, including the current shortcomings of empirical scaling approaches and the lack of suitable mechanistic approaches.
bioavailability; hypodermis; lymphatic uptake; subcutaneous; therapeutic proteins
We conducted a pharmacokinetic (PK) study of mitoxantrone (Novantrone®), a clinically well-established anticancer agent, in mice and developed a mechanism-based PBPK (physiologically based pharmacokinetic) model to describe its disposition. Mitoxantrone concentrations in plasma and six organs (lung, heart, liver, kidney, spleen, and brain) were determined after a 5 mg/kg i.v. dose. We evaluated three different PBPK models in order to characterize our experimental data: model 1 containing Kp values, model 2 incorporating a deep binding compartment, and model 3 incorporating binding of mitoxantrone to DNA and protein. Among the three models, only model 3 with DNA and protein binding captured all the experimental data well. The estimated binding affinity for DNA (KDNA) and protein (Kmacro) were 0.0013 and 1.44 μM, respectively. Predicted plasma and tissue AUC values differed from observed values by <19 %, except for heart (60 %). Model 3 was further used to simulate plasma mitoxantrone concentrations in humans for a 12-mg/m2 dose, using human physiological parameters. The simulated results generally agreed with the observed time course of mitoxantrone plasma concentrations in patients after a standard dose of 12 mg/m2. In summary, we reported for the first time a mechanism-based PBPK model of mitoxantrone incorporating macromolecule binding which may have clinical applicability in optimizing clinical therapy. Since mitoxantrone is a substrate of the efflux transporters ABCG2 and ABCB1, the incorporation of efflux transporters may also be necessary to characterize the data obtained in low-dose studies.
DNA binding; human scale-up; mitoxantrone; physiologically based pharmacokinetic model
Our objective was to determine the pharmacokinetics, bioavailability and lymph node uptake of the monoclonal antibody bevacizumab, labeled with the near-infrared (IR) dye 800CW, after intravenous (IV) and subcutaneous (SC) administration in mice. Fluorescence imaging and enzyme-linked immunosorbent assay (ELISA) assays were developed and validated to measure the concentration of bevacizumab in plasma. The bevacizumab–IRDye conjugate remained predominantly intact in plasma and in lymph node homogenate samples over a 24-h period, as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis and size exclusion chromatography. The plasma concentration vs. time plots obtained by fluorescence and ELISA measurements were similar; however, unlike ELISA, fluorescent imaging was only able to quantitate concentrations for 24 h after administration. At a low dose of 0.45 mg/kg, the plasma clearance of bevacizumab was 6.96 mL/h/kg after IV administration; this clearance is higher than that reported after higher doses. Half-lives of bevacizumab after SC and IV administration were 4.6 and 3.9 days, respectively. After SC administration, bevacizumab–IRDye800CW was present in the axillary lymph nodes that drain the SC site; lymph node uptake of bevacizumab–IRDye 800CW was negligible after IV administration. Bevacizumab exhibited complete bioavailability after SC administration. Using a compartmental pharmacokinetic model, the fraction absorbed through the lymphatics after SC administration was estimated to be about 1%. This is the first report evaluating the use of fluorescent imaging to determine the pharmacokinetics, lymphatic uptake, and bioavailability of a near-infrared dye-labeled antibody conjugate.
Electronic supplementary material
The online version of this article (doi:10.1208/s12248-012-9342-9) contains supplementary material, which is available to authorized users.
bevacizumab; bioavailability; fluorescence imaging; lymphatic absorption; pharmacokinetics
γ-Hydroxybutyric acid (GHB), a drug of abuse, demonstrates complex toxicokinetics with capacity-limited metabolism and active renal reabsorption. The objectives of the present study were to conduct a local sensitivity analysis of a mechanistic model for the active renal reabsorption of GHB and to use the results to inform the design of future studies aimed at developing therapeutic strategies for treating GHB overdoses. A local sensitivity analysis was used to assess the influence of parameter perturbations on model outputs (plasma concentrations and urinary excretion of GHB). Further, a sensitivity index was calculated for each perturbed parameter to assess the specific segments of the time course that are critical to parameter estimation. Model outputs were simulated for rats dosed with 200, 400, 600, and 1,000 mg/kg GHB intravenously and individual parameters were perturbed by two-, five-, and tenfold higher and lower than the nominal value. Model outputs were sensitive to perturbations in clearance and volume parameters. In contrast, model outputs were found to be insensitive to changes in distributional parameters suggesting that additional tissue distribution data is required. Based on the sensitivity analysis the 1,000-mg/kg GHB dose can be eliminated from future studies as the parameters can be adequately estimated from the lower doses. To further validate the use of this model, dose-specific sampling schedules were designed based on model predictions for doses of 600 and 1,500 mg/kg. These sampling schedules were able to adequately capture the inflection point and terminal elimination phase of the plasma concentration–time profiles obtained.
active renal reabsorption; gamma-hydroxybutyric acid; monocarboxylate transporters; sensitivity analysis
γ-Hydroxybutyric acid (GHB), a drug of abuse, exhibits saturable renal clearance and capacity-limited metabolism. The objectives of this study were to construct a mechanistic toxicokinetic (TK) model describing saturable renal reabsorption and capacity-limited metabolism of GHB and to predict the effects of inhibition of renal reabsorption on GHB TK in the plasma and urine. GHB was administered by iv bolus (200–1,000 mg/kg) to male Sprague-Dawley rats and plasma and urine samples were collected for up to 6 h post-dose. GHB concentrations were determined by LC/MS/MS. GHB plasma concentration and urinary excretion were well-described by a TK model incorporating plasma and kidney compartments, along with two tissue and two ultrafiltrate compartments. The estimate of the Michaelis-Menten constant for renal reabsorption (Km,R) was 0.46 mg/ml which is consistent with in vitro estimates of monocarboxylate transporter (MCT)-mediated uptake of GHB (0.48 mg/ml). Simulation studies assessing inhibition of renal reabsorption of GHB demonstrated increased time-averaged renal clearance and GHB plasma AUC, independent of the inhibition mechanism assessed. Co-administration of GHB (600 mg/kg iv) and l-lactate (330 mg/kg iv bolus plus 121 mg/kg/h iv infusion), a known inhibitor of MCTs, resulted in a significant decrease in GHB plasma AUC and an increase in time-averaged renal clearance, consistent with the model simulations. These results suggest that inhibition of renal reabsorption of GHB is a viable therapeutic strategy for the treatment of GHB overdoses. Furthermore, the mechanistic TK model provides a useful in silico tool for the evaluation of potential therapeutic strategies.
gamma-hydroxybutyrate; kidney reabsorption; pharmacokinetic model; renal clearance; toxicokinetics
Breast cancer resistance protein (ABCG2), the newest ABC transporter, was discovered independently by three groups in the late 1990s. ABCG2 is widely distributed in the body with expression in the brain, intestine, and liver, among others. ABCG2 plays an important role by effluxing drugs at the blood–brain, blood–testis, and maternal–fetal barriers and in the efflux of xenobiotics at the small intestine and kidney proximal tubule brush border and liver canalicular membranes. ABCG2 transports a wide variety of substrates including HMG-CoA reductase inhibitors, antibiotics, and many anticancer agents and is one contributor to multidrug resistance in cancer cells. Quantitative structure–activity relationship (QSAR) models and structure–activity relationships (SARs) are often employed to predict ABCG2 substrates and inhibitors prior to in vitro and in vivo studies. QSAR models correlate in vivo biological activity to physicochemical properties of compounds while SARs attempt to explain chemical moieties or structural features that contribute to or are detrimental to the biological activity. Most ABCG2 datasets available for in silico modeling are comprised of congeneric series of compounds; the results from one series usually cannot be applied to another series of compounds. This review will focus on in silico models in the literature used for the prediction of ABCG2 substrates and inhibitors.
ABC transporter; ABCG2; breast cancer resistance protein; quantitative structure–activity relationships; structure–activity relationships
The aims were (1) to evaluate the molecular weight (MW) dependence of biliary excretion and (2) to develop quantitative structure–pharmacokinetic relationships (QSPKR) to predict biliary clearance (CLb) and percentage of administered dose excreted in bile as parent drug (PDb) in rats and humans. CLb and PDb data were collected from the literature for rats and humans. Receiver operating characteristic curve analysis was utilized to determine whether a MW threshold exists for PDb. Stepwise multiple linear regression (MLR) was used to derive QSPKR models. The predictive performance of the models was evaluated by internal validation using the leave-one-out method and external test groups. A MW threshold of 400 Da was determined for PDb for anions in rats, while 475 Da was the cutoff for anions in humans. MW thresholds were not present for cations or cations/neutral compounds in either rats or humans. The QSPKR model for human CLb showed a significant correlation (R2 = 0.819) with good prediction performance (Q2 = 0.722). The model was further assessed using a test group, yielding a geometric mean fold-error of 2.68. QSPKR models with significant correlation and good predictability were also developed for CLb in rats and PDb data for anions or cation/neutral compounds in rats and humans. Both CLb and PDb data were further evaluated for subsets of MRP2 or P-glycoprotein substrates, and significant relationships were derived. QSPKR models were successfully developed for biliary excretion of non-congeneric compounds in rats and humans, providing a quantitative prediction of biliary clearance of compounds.
biliary clearance; humans; in silico; molecular weight cutoff; molecular volume; QSPKR; rats
Monocarboxylate transporter 1 (MCT1) has been previously reported as an important determinant of the renal reabsorption of the drug of abuse, γ-hydroxybutyrate (GHB). Luteolin is a potent MCT1 inhibitor, inhibiting the uptake of GHB with an IC50 of 0.41 μM in MCT1-transfected MDA-MB231 cells. The objectives of this study were to characterize the effects of luteolin on GHB pharmacokinetics and pharmacodynamics in rats, and to investigate the mechanism of the interaction using model-fitting methods. GHB (400 and 1,000 mg/kg) and luteolin (0, 4 and 10 mg/kg) were administered to rats via iv bolus doses. The plasma or urine concentrations of luteolin and GHB were determined by HPLC and LC/MS/MS, respectively. The pharmacodynamic parameter sleep time in rats after GHB administration was recorded. A pharmacokinetic model containing capacity-limited renal reabsorption and metabolic clearance was constructed to characterize the in vivo interaction. Luteolin significantly decreased the plasma concentration and AUC, and increased the total and renal clearances of GHB. Moreover, luteolin significantly shortened the duration of GHB (1,000 mg/kg)-induced sleep in rats (161 ± 16, 131 ± 14 and 121 ± 5 min for control, luteolin 4 and 10 mg/kg groups, respectively, p < 0.01). An uncompetitive inhibition model, with an inhibition constant of 1.1 μM, best described the in vivo pharmacokinetic interaction. The results of this study indicated that luteolin significantly altered the pharmacokinetics of GHB by inhibiting its MCT1-mediated transport. The interaction between luteolin and GHB may offer a potential clinical detoxification strategy to treat GHB overdoses.
γ-hydroxybutyrate; luteolin; MCT; pharmacokinetic interactions
The American Association of Pharmaceutical Scientists (AAPS) covers the full range of areas of expertise associated with the resolution of concerns pertaining to drugs and drug products. This editorial highlights the initiatives, issues, and challenges that are the forefront of the pharmaceutical sciences in 2007. It also provides an overview of how these difficult questions are being addressed through the programs and events associated with the AAPS 2007 Annual Meeting that will be held at the San Diego, California, Convention Center from November 11 to 15, 2007.
dose predictions; product design; product quality control; population kinetics; dose individualization; regulatory sciences; pharmacostatistics; process analytical technology; medical imagining; quantitative pharmacology; dissolution; biotechnology
Biochanin A(BCA) is a dietary isoflavone present in legumes, most notably red clover, and in many herbal dietary supplements. BCA has been reported to have chemopreventive properties and is metabolized to the isoflavone genistein (GEN), BCA conjugates, and GEN conjugates. The metabolites may contribute to the chemopreventive effects of BCA. The absorption, metabolism, and disposition of BCA have not been determined in rats. Our objective was to evaluate the pharmacokinetics and metabolism of BCA in rats. Male Sprague-Dawley rats were administered BCA by intravenous injection (1 and 5 mg/kg), by intraperitoneal injection (5 and 50 mg/kg), and orally (5 and 50 mg/kg). Plasma and bile samples were enzymatically hydrolyzed in vitro to determine conjugate concentrations for BCA and GEN. Equilibrium dialysis was used to determine protein binding. The BCA and GEN concentrations in plasma, urine, and bile were determined by liquid chromatography-tandem mass spectrometry (LC/MS/MS). The pharmacokinetic parameters of BCA were analyzed by noncompartmental analysis. Significant levels of BCA conjugates and GEN conjugates were detected in plasma and bile. Both BCA and GEN were found to have a high clearance and a large apparent volume of distribution; the bioavailability of both was poor (<4%). Reentry peaks were evident after oral administration of both BCA and GEN, suggesting enterohepatic cycling. The free fraction of BCA in rat plasma was 1.5%. A2-compartment model that included both linear and nonlinear clearance terms and enterohepatic recirculation best described the plasma data. This represents the first evaluation of the dose-dependent pharmacokinetics and metabolism of BCA in rats.
Biochanin A; pharmacokinetics; intraperitoneal administration; enterohepatic recirculation; rat; genistein