Drug Metabolism and Disposition最新文献

The drug candidate, 2-methoxy-N-[3-[4-[3-methyl-4-[(6-methyl-3-pyridinyl)oxy]anilino]-6-quinazolinyl]prop-2-enyl]acetamide (CP-724,714), was discontinued because of hepatotoxicity observed in clinical studies. CP-724,714 is a substrate of aldehyde oxidase (AO) found in the human liver cytosol. CP-724,714 metabolization by AO in cryopreserved human hepatocytes generates several oxidative metabolites, including (E)-N-(3-(2-hydroxy-4-(3-methyl-4-(6-methylpyridin-3-yloxy)phenylamino)quinazolin-6-yl)allyl)-2-methoxyacetamide (CP-724,714-AOM). However, the structure of CP-724,714-AOM has not been identified. Therefore, we aimed to identify the structure of CP-724,714-AOM, determine the propensity of CP-724,714 and CP-724,714-AOM for toxic effects, and understand the underlying hepatotoxic mechanism in humans. A synthesized oxidized CP-724,714, identified as CP-724,714-AOM, was consistent with the AO metabolite of CP-724,714 generated in the human liver cytosol. The enzymatic kinetic parameters of CP-724,714 were calculated as a K_m value of 9.17 ± 0.70 μM and a V_max value of 3.57 ± 0.10 pmol/min/mg of human liver cytosol proteins, assessed by the production of CP-724,714-AOM. CP-724,714 showed a weak inhibitory effect on the bile salt export pump without inhibiting multidrug resistance protein 2, whereas CP-724,714-AOM showed no inhibitory effect. The trapping assay showed that both compounds formed reactive metabolites in the human liver microsomes. Moreover, the inflammasome activation potentials of the compounds were observed in HepaRG cells, which can also generate CP-724,714-AOM from CP-724,714. Thus, our findings show that confirming AO susceptibility at an early stage of drug development is crucial for understanding the potential risks of AO metabolism in terms of pharmacokinetics and toxicity. SIGNIFICANCE STATEMENT: The structure of the aldehyde oxidase metabolite of 2-methoxy-N-[3-[4-[3-methyl-4-[(6-methyl-3-pyridinyl)oxy]anilino]-6-quinazolinyl]prop-2-enyl]acetamide (CP-724,714) was identified using a synthetic standard, and its kinetic parameters in the human liver cytosol were determined. Reactive metabolite formation and inflammasome activation by CP-724,714 and (E)-N-(3-(2-hydroxy-4-(3-methyl-4-(6-methylpyridin-3-yloxy)phenylamino)quinazolin-6-yl)allyl)-2-methoxyacetamide were observed.

This study investigated the effects of indoxyl sulfate (IS), an endogenous metabolite and uremic toxin, on arsenic trioxide pharmacokinetics in acute promyelocytic leukemia patients with varying renal function. Plasma IS levels demonstrated a significant positive correlation with monomethylarsonic acid and dimethylarsinic acid concentrations in patients (P < .0001). In adenine-induced renally impaired rats, IS similarly correlated with elevated plasma inorganic arsenic (iAs), monomethylarsonic acid, and dimethylarsinic acid levels. Protein expression analysis indicated a downregulation of renal aquaporin (AQP) 7 and AQP3. In vitro studies confirmed that IS selectively inhibits AQP7 expression (62.1% reduction at 100 μM) through aryl hydrocarbon receptor activation in human embryonic kidney 293T cells, while AQP3 remained unaffected. Collectively, IS increases plasma arsenic concentration in renally impaired acute promyelocytic leukemia patients via aryl hydrocarbon receptor-mediated suppression of renal AQP7. SIGNIFICANCE STATEMENT: This study reveals that indoxyl sulfate inhibits renal aquaporin 7 via aryl hydrocarbon receptor activation, increasing plasma arsenic in arsenic trioxide-treated acute promyelocytic leukemia patients with renal impairment. As the first demonstration of this mechanism, to our knowledge, it provides crucial insights for optimizing therapy and reducing toxicity risks.

COVID-19 remains a significant health threat, particularly to people over the age of 65 with existing comorbidities like diabetes, hypertension, cancer, and viral infections. Despite expedited drug approvals, drug-drug interaction profiles for COVID-19 antiviral drugs have not yet been completely defined. The antiviral drugs remdesivir and molnupiravir are ester prodrugs with carboxylesterases (CES) playing a critical role in their bioactivation. In this study, we investigated the effect of COVID-19 antiviral drugs on CES hydrolysis activity. Of the 3 drugs tested, remdesivir inhibited 50% of CES2 activity in a nanomolar concentration range. Furthermore, time-dependent inhibition of CES2 activity by remdesivir was identified with an IC₅₀ shift of nearly 3-fold from 0.188 μM after 5-minute preincubation to 0.068 μM following 40-minute preincubation. Remdesivir inactivation of CES2 was characterized by a k_inact/K_I value of 1.6 × 10³ M^-1.s^-1. Through equilibrium dialysis and substrate protection experiments, we were able to further substantiate previous findings of the irreversible CES2 inhibition by remdesivir. Finally, in silico docking analysis of remdesivir to CES2 supported the proposed mechanism for covalent modification of the CES2 active site involving the catalytic triad serine via the phosphate group of remdesivir. SIGNIFICANCE STATEMENT: This study provides insights into human carboxylesterase inhibition by COVID-19 drugs and these findings demonstrate the underlying inhibition mechanism by remdesivir of recombinant human carboxylesterase 2.

Three-dimensional (3D) cultures of primary human hepatocytes (3D PHH) are successfully used to reduce and replace the use of animal experiments in biomedical research. Yet, the initial formation of 3D PHH is highly dependent on the supplementation with FBS. However, the molecular composition of FBS and its effects on cultured cells are poorly understood. Moreover, FBS is prone to batch-to-batch variation, immunogenic risk, and lack of adherence to the replacement, refinement, and reduction of animal experiments. Here, we demonstrate that FBS can be fully replaced by animal-free substitutes, thus facilitating fully chemically defined and animal serum-free 3D PHH cultures. Specifically, we combined a previously developed animal-free substitute cocktail with a normoglycemic (5.5 mM glucose and 0.58 ng/mL insulin) chemically defined culture medium. Morphological and viability evaluations, along with global proteomics data, demonstrated that serum-free cultured 3D PHH have comparable viability and functional performance of cytochrome P450s, rendering this medium useful for long-term studies and in vitro absorption, distribution, metabolism, excretion, and toxicity applications. This study marks a significant advancement in the development of animal serum-free culture conditions for primary human cell cultures, paving the way for more reliable and ethical in vitro studies. SIGNIFICANCE STATEMENT: Most in vitro cell models rely on FBS. However, the use of FBS leads to inconsistent experimental results and raises serious ethical concerns. In this study, a chemically defined animal product-free cell culture medium with physiologically relevant levels of key hormones and nutrients for liver spheroid cultures was developed and evaluated. This study marks a significant advancement in the development of animal serum-free culture conditions for primary human cell cultures used in drug disposition studies.

Cytochrome P450 (P450)-mediated drug-drug interactions (DDIs) are responsible for most adverse drug interactions, and occur when 2 concurrently administered drugs inhibit, upregulate, or are substrates of the same target enzyme. A machine learning approach enables the detection of DDIs with rarely used drugs, as well as newly approved drugs. To facilitate this, we present a framework for predicting DDIs by first predicting P450 interactions for both drugs, generating a fingerprint based on the predictions in addition to the molecular structures of the drugs, and training a machine learning model to predict the overall interaction. After optimization, the model detected potential DDIs with 85% accuracy, representing an improvement over a DDI-only model (ie, a model trained on structure-based fingerprints without supporting P450 model predictions). We also present a corresponding adverse outcome pathway to allow for increased model explainability through visualizing each predicted P450 interaction, further enhancing its real-world applicability. Finally, we show the importance of the model applicability domain to DDI models by demonstrating how the performance of our model degrades as the inference set becomes dissimilar to the training data. SIGNIFICANCE STATEMENT: Polypharmacotherapy (especially in older populations) results in more drugs prescribed concurrently, creating an increased risk of drug-drug interactions and adverse drug reactions. Computational tools to predict potential drug-drug interactions could accurately aid in reducing risk for the patient and be used in the early stages of drug design to avoid such undesirable molecular interactions.

Hydromorphone is a highly potent opioid used to treat severe chronic pain. It is metabolized primarily by UDP-glucuronosyltransferase (UGT)2B7 to form the inactive hydromorphone-3-glucuronide. Given that previous studies have shown that the major cannabinoids, Δ⁹-tetrahydrocannabinol (THC) and cannabidiol (CBD), inhibit several UGT enzymes, the objective of the present study was to determine the inhibitory potential of major cannabinoids and their metabolites on UGT-mediated hydromorphone metabolism. To evaluate the potential for cannabis-induced drug interactions, cannabinoids and their metabolites were screened as potential inhibitors against hydromorphone glucuronidation in pooled human liver microsomes and microsomes from cells overexpressing recombinant UGT2B7. IC₅₀ values were determined for cannabinoids that inhibited hydromorphone glucuronidation by 50% and K_i values for those that exhibited an IC₅₀ < 100 μM in human liver microsomes. Potent inhibition of hydromorphone metabolism was observed for THC, 11-hydroxy (OH)-THC, CBD, and 7-OH-CBD, with K_i values ranging from 0.068 to 1.01 μM after correction for nonspecific cannabinoid binding. Differences in inhibition were observed for the UGT2B7^268Tyr variant compared with the wildtype UGTB7^268His isoform for several cannabinoids. Static modeling indicated that THC, 11-OH-THC, CBD, and 7-OH-CBD would result in drug interactions in vivo after inhalation and oral consumption of THC and CBD (>1.25-fold increase in hydromorphone exposure), with physiologically based pharmacokinetic predictive models indicating that CBD would cause a 20%-30% increase in hydromorphone exposure in healthy and cirrhotic individuals. These data suggest that major cannabinoids such as CBD will cause moderate drug-drug interactions with hydromorphone in humans. SIGNIFICANCE STATEMENT: This study indicates that major cannabinoids and their metabolites found in the plasma of cannabis users inhibit UGT2B7-mediated hydromorphone metabolism in vitro. It further demonstrates the potential for in vivo inhibition of hydromorphone metabolism by cannabinoids and their metabolites, indicating the potential for drug-drug interactions upon concomitant use of hydromorphone and cannabis or hydromorphone together with individual cannabinoids like Δ⁹-tetrahydrocannabinol and cannabidiol.

Cyclosporine A (CsA) is a widely used immunosuppressant for posttransplantation and autoimmune diseases, but its clinical application is limited by severe hepatorenal toxicity. Krüppel-like factor 15 (KLF15), a key regulator of xenobiotic metabolism, has been shown to modulate the metabolism of acetaminophen and rifampicin and play a role in the associated drug-induced liver toxicity. However, its role in CsA metabolism and CsA-induced hepatorenal injury remains unclear. This study aimed to investigate whether hepatic KLF15 regulates CsA metabolism and serves as a potential therapeutic target for mitigating CsA-induced hepatorenal toxicity. KLF15 broadly suppressed the CsA detoxification pathways by inhibiting key metabolic enzymes and transporters. Inhibition of hepatic KLF15 enhanced CsA detoxification, reduced CsA accumulation, and prevented hepatorenal injury. Notably, both AAV-shKlf15 and PXR agonist treatment demonstrated protective effects even after CsA-induced organ damage had occurred. These findings highlight KLF15 as a critical regulator of CsA metabolism and a promising therapeutic target for preventing or treating CsA-induced hepatorenal toxicity. The study provides preclinical evidence supporting further exploration of KLF15 modulation in clinical settings. SIGNIFICANCE STATEMENT: Hepatic Krüppel-like factor 15 is critical in regulating the metabolism of acetaminophen, rifampicin, and cyclosporine A, drugs used for pain relief, antibiotics, and immune system regulation, respectively. It may serve as a potential therapeutic target for liver and kidney damage induced by these drugs.

SAF-189s is a promising highly selective, brain-penetrant, next-generation inhibitor of anaplastic lymphoma kinase (ALK) and c-ROS proto-oncogene 1 (ROS1). The pharmacokinetics, mass balance, and metabolism of SAF-189s were measured in 6 healthy Chinese male participants after receiving a single oral dose of 160 mg [¹⁴C]SAF-189s (150 μCi). SAF-189s was rapidly absorbed with a median T_max of 4.0 hours. The arithmetic mean half-life of total radioactivity in plasma was approximately 32.1 hours. The ratio of mean total drug-related substance concentration in whole blood to that in plasma (B/P_AUC) was 3.06, indicating that the drug was predominantly distributed in blood cells. After 336 hours of drug administration, the average cumulative excretion of radioactivity accounted for 96.98% of the total dose, with 6.24% of the drug excreted in urine and 90.74% of the drug excreted in feces. In total, 14 metabolites were identified. SAF-189s was the predominant component in plasma but was scarcely detected in urine and feces. Oxidative metabolism mediated by CYP3A4 was determined to be the primary metabolic pathway for SAF-189s, with the isopropyl group being the most susceptible metabolizing site. M543 was identified as the main oxidative metabolite of SAF-189s in humans, and its production was likely affected by both CYP3A and intestinal microbiota. After a single oral dose of [¹⁴C]SAF-189s, SAF-189s and its principal metabolites were primarily excreted via feces. The main metabolic pathway was oxidation, likely catalyzed by both CYP3A and intestinal microbiota. SIGNIFICANCE STATEMENT: This study investigated the absorption and disposition of SAF-189s, a promising next-generation inhibitor of ALK/ROS1 administered for the treatment of ALK+/ROS1+ non-small cell lung cancer. The results demonstrated that SAF-189s and its metabolites were primarily excreted via feces, with metabolism likely mediated by both the cytochrome P450 system and gut microbiota. These findings provide essential pharmacokinetic and safety data, encourage further studies on drug interactions and dose adjustments, and support the involvement of gut microbiota, thereby guiding future research.

Hypogonadism, characterized by low testosterone blood levels, affects 3%-5% of males worldwide. Oral testosterone undecanoate (TU) is emerging as a key route of administration due to its better ease of administration; however, it suffers from variable pharmacokinetics and pharmacodynamics. The variability is majorly attributed to intestinal glucuronidation of testosterone to its hydrophilic metabolite, testosterone glucuronide (TG), formed by the polymorphic uridine 5'-diphospho-glucuronosyltransferase 2B17 (UGT2B17). This study investigated the potential of curcumin, as a UGT2B17 inhibitor, to enhance TU bioavailability using a series of in vitro and in vivo studies. In human intestinal microsomes, curcumin inhibited UGT2B17 with an IC₅₀ of 58 μM. In LS180 cells, a human intestinal cell line, curcumin reduced TG formation dose-dependently at 10, 25, and 100 μM, and also inhibited the formation of androstenedione at 100 μM. In primary human enterocytes, curcumin (100 μM) significantly reduced TG and androstenedione formation by ∼50%. A pilot crossover clinical study compared testosterone pharmacokinetics when TU was coadministered with curcumin in men with experimental hypogonadism. The plasma concentration-time area under the curve (AUC_1-6h) and the peak plasma concentration (C_max) for testosterone, representing the absorption phase, significantly increased by 50% and 80%, respectively, when TU was administered with curcumin. These findings suggest that curcumin, a UGT2B17 inhibitor, has the potential to enhance the bioavailability of oral TU. Further clinical investigations with higher doses of curcumin and with other UGT2B17 inhibitors in diverse populations are warranted to validate the UGT2B17 inhibition approach for enhancing oral testosterone replacement therapy. SIGNIFICANCE STATEMENT: Hypogonadism affects a significant portion of the male population. Oral testosterone undecanoate is recommended to treat hypogonadism, but shows variable pharmacokinetics and pharmacodynamics. This study evaluated curcumin as a potential enhancer of testosterone undecanoate bioavailability by inhibiting intestinal UGT2B17, the key enzyme mediating testosterone glucuronidation. In vitro and pilot clinical studies showed that curcumin can inhibit testosterone metabolism and enhance its systemic levels. These findings suggest that UGT2B17 inhibition by curcumin or other UGT2B17 inhibitors could be a promising approach to improve oral testosterone bioavailability.