Pharmacokinetic analysis of antiviral drug ritonavir across the blood-brain barrier and its interaction with Scutellaria baicalensis using multisite microdialysis in rats.

Ritonavir, an excellent inhibitor of CYP3A4, has recently been combined with nirmatrelvir to form Paxlovid for the treatment of severe acute respiratory syndrome coronavirus 2 infections. The root of Scutellaria baicalensis Georgi (S. baicalensis), a traditional Chinese medicinal (TCM) herb commonly used to treat heat/inflammation in the lung and digestive tracts, which are major organs targeted by viral infections, contains flavones that can influence the CYP3A metabolism pathway. To investigate the ability of ritonavir to cross the bloodbrain barrier (BBB) and its potential herb-drug interactions with an equivalent TCM clinical dose of S. baicalensis, multisite microdialysis coupled with an LCMS/MS system was developed using rat model. Pretreatment with S. baicalensis extract for 5 days, which contains less flavones than those used in previous studies, had a significant influence on ritonavir, resulting in a 2-fold increase in the total concentration of flavones in the blood and brain. Treatment also boosted the maximum blood concentration of flavones by 1.5-fold and the maximum brain concentration of flavones by 2-fold, all the while exerting no noticeable influence on the transfer ratio across the bloodbrain barrier. These experimental results demonstrated that the use of a typical traditional Chinese medicinal dose of S. baicalensis is sufficient to influence the metabolic pathway and synergistically increase the concentration of ritonavir in rats.

Pharmacokinetic analysis of placental transfer of ritonavir as a component of paxlovid using microdialysis in pregnant rats.

Background: Ritonavir is one of the most potent CYP3A4 inhibitor currently on the market, and is often used together with other antiviral drugs to increase their bioavailability and efficacy. Paxlovid, consisting of nirmatrelvir and ritonavir, was approved for the treatment of COVID-19. As previous studies regarding the use of ritonavir during pregnancy were limited to ex-vivo experiments and systemic safety data, to fully explore the detailed pharmacokinetics of ritonavir in pregnant rats' blood and conceptus, an analytical method consisted of multi-microdialysis coupled with UHPLC-MS/MS were developed to analyze the pharmacokinetics of ritonavir, both as a component of Paxlovid and by itself. 17 days pregnant female Sprague-Dawley rats were randomly split into three experimental group: normal dosage of ritonavir alone (7 mg kg-1), normal dosage of Paxlovid (ritonavir 7 mg kg-1 + nirmatrelvir 15 mg kg-1), and 3× dosage of ritonavir (21 mg kg-1). Results: 3× dosage of ritonavir produced a more than 3× increase in rats' blood and placenta. Transfer rate of ritonavir to the placenta, amniotic fluid, and fetus were determined to be 20.7%, 13.8%, and 4.7% respectively. Concentration of ritonavir in the placenta, amniotic fluid, and fetus did not significantly go down after 8 h. Significance: Overall, ritonavir's metabolism was not influenced by the presence of nirmatrelvir in pregnant rats. A 3× increase in dosage produced a concentration of roughly 4×, most likely a result of ritonavir's auto-inhibition effect on cytochrome P450 proteins. Accumulation of ritonavir is possible in placenta, amniotic fluid, and fetus.

The clinical application of traditional Chinese medicine NRICM101 in hospitalized patients with COVID-19.

BACKGROUND: The aim of this study was to assess the efficacy and safety of NRICM101 in hospitalized patients with COVID-19. RESEARCH DESIGN AND METHODS: We conducted a retrospective study from 20 April 2021 to 8 July 2021, and evaluated the safety and outcomes (mortality, hospital stay, mechanical ventilation, oxygen support, diarrhea, serum potassium) in COVID-19 patients. Propensity score matching at a 1:2 ratio was performed to reduce confounding factors. RESULTS: A total of 201 patients were analyzed. The experimental group (n = 67) received NRICM101 and standard care, while the control group (n = 134) received standard care alone. No significant differences were observed in mortality (10.4% vs. 14.2%), intubation (13.8% vs. 11%), time to intubation (10 vs. 11 days), mechanical ventilation days (0 vs. 9 days), or oxygen support duration (6 vs. 5 days). However, the experimental group had a shorter length of hospitalization (odds ratio = 0.12, p = 0.043) and fewer mechanical ventilation days (odds ratio = 0.068, p = 0.008) in initially severe cases, along with an increased diarrhea risk (p = 0.035). CONCLUSION: NRICM101 did not reduce in-hospital mortality. However, it shortened the length of hospitalization and reduced mechanical ventilation days in initially severe cases. Further investigation is needed.

Transporter modulation of molnupiravir and its metabolite ß-D-N4-hydroxycytidine across the blood-brain barrier in a rat.

BACKGROUND: The antiviral drug molnupiravir is an orally bioavailable prodrug of the nucleoside analog ß-D-N4-hydroxycytidine (NHC), which is used to treat coronavirus disease 2019 (COVID-19). However, there is very little information on the barrier distribution of molnupiravir. Our hypothesis is that molnupiravir and NHC can penetrate the blood‒brain barrier (BBB) into brain tissue and that nucleoside transporters (equilibrative nucleoside transporters; ENT and concentrative nucleoside transporters; CNT) can modulate this process. METHODS: To investigate the mechanism of molnupiravir transport through the BBB, multiple microdialyses coupled to a validated ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC‒MS/MS) was developed to monitor dialysates, and nitrobenzylthioinosine (NBMPR; an inhibitor of ENT) was administered concomitantly with molnupiravir (100 mg/kg, i.v.) in the male rat. RESULTS: Here, we show that molnupiravir is rapidly metabolized to NHC in the blood and crossed the BBB in 20 min. Furthermore, when NBMPR is concomitantly administered to inhibit efflux, the concentrations of molnupiravir and NHC in the brain increased significantly. CONCLUSIONS: In summary, molnupiravir rapidly transforms into NHC and crosses the BBB and reaches the brain at approximately 0.3-0.8% of the blood‒brain ratio. The maximum concentration of NHC in the blood and brain is above the average half maximal inhibitory concentration (IC50) of the drug required to treat severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, suggesting a therapeutic effect. The penetration of NHC is modulated by NBMPR. These findings provide constructive information on brain disorders in clinical patients with COVID-19.

Due to the global pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome (SARS-CoV-2), molnupiravir is used orally to treat COVID-19 with emergency use authorization. However, it is not well understood whether molnupiravir and its active component can cross the blood­brain barrier. The aim of the study was to develop an experimental mouse model to monitor the journey of molnupiravir and its active component through the bloodstream and eventually into the brain. Our experimental data suggest that a therapeutically useful amount of molnupiravir crosses from the bloodstream into the brain.

Transfer and biotransformation of the COVID-19 prodrug molnupiravir and its metabolite ß-D-N4-hydroxycytidine across the blood-placenta barrier.

BACKGROUND: Molnupiravir is an orally bioavailable prodrug of the nucleoside analogue ß-D-N4-hydroxycytidine (NHC) and is used to treat coronavirus disease 2019 (COVID-19). However, the pharmacokinetics and transplacental transfer of molnupiravir in pregnant women are still not well understood. In the present study, we investigated the hypothesis that molnupiravir and NHC cross the blood-placenta barrier into the fetus. METHODS: A multisite microdialysis coupled with a validated ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC‒MS/MS) system was developed to monitor the dialysate levels of molnupiravir and NHC in maternal rat blood and conceptus (the collective term for the fetus, placenta, and amniotic fluid). Molnupiravir was administered intravenously (100 mg/kg, i.v.) on gestational day 16. To investigate the mechanism of transport of molnupiravir across the blood-placenta barrier, we coadministered nitrobenzylthioinosine (NBMPR, 10 mg/kg, i.v.) to inhibit equilibrative nucleoside transporter (ENT). FINDINGS: We report that molnupiravir is rapidly metabolized to NHC and then rapidly transformed in the fetus, placenta, amniotic fluid, and maternal blood. Our pharmacokinetics analysis revealed that the area under the concentration curve (AUC) for the mother-to-fetus ratio (AUCfetus/AUCblood) of NHC was 0.29 ± 0.11. Further, we demonstrated that the transport of NHC in the placenta may not be subject to modulation by the ENT. INTERPRETATION: Our results show that NHC is the predominant bioactive metabolite of molnupiravir and rapidly crosses the blood-placenta barrier in pregnant rats. The NHC concentration in maternal blood and conceptus was above the average median inhibitory concentration (IC50) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), suggesting a therapeutic effect. These findings support the use of molnupiravir in pregnant patients infected with COVID. FUNDING: This study was supported in part by research grants from the National Science and Technology Council of Taiwan (NSTC 111-2113-M-A49-018 and NSTC 112-2321-B-A49-005).

Biotransformation and brain distribution of the anti-COVID-19 drug molnupiravir and herb-drug pharmacokinetic interactions between the herbal extract Scutellaria formula-NRICM101.

The aim of this study was to explore the effects of herbal drug pharmacokinetic interactions on the biotransformation of molnupiravir and its metabolite ß-D-N4-hydroxycytidine (NHC) in the blood and brain. To investigate the biotransformation mechanism, a carboxylesterase inhibitor, bis(4-nitrophenyl)phosphate (BNPP), was administered. Not only molnupiravir but also the herbal medicine Scutellaria formula-NRICM101 is potentially affected by coadministration with molnupiravir. However, the herb-drug interaction between molnupiravir and the Scutellaria formula-NRICM101 has not yet been investigated. We hypothesized that the complex bioactive herbal ingredients in the extract of the Scutellaria formula-NRICM101, the biotransformation and penetration of the bloodbrain barrier of molnupiravir are altered by inhibition of carboxylesterase. To monitor the analytes, ultrahigh-performance liquid chromatography tandem mass spectrometry (UHPLCMS/MS) coupled with the microdialysis method was developed. Based on the dose transfer from humans to rats, a dose of molnupiravir (100 mg/kg, i.v.), molnupiravir (100 mg/kg, i.v.) + BNPP (50 mg/kg, i.v.), and molnupiravir (100 mg/kg, i.v.) + the Scutellaria formula-NRICM101 extract (1.27 g/kg, per day, for 5 consecutive days) were administered. The results showed that molnupiravir was rapidly metabolized to NHC and penetrated into the brain striatum. However, when concomitant with BNPP, NHC was suppressed, and molnupiravir was enhanced. The blood-to-brain penetration ratios were 2% and 6%, respectively. In summary, the extract of the Scutellaria formula-NRICM101 provides a pharmacological effect similar to that of the carboxylesterase inhibitor to suppress NHC in the blood, and the brain penetration ratio was increased, but the concentration is also higher than the effective concentration in the blood and brain.

Simultaneous toxicokinetic studies of aristolochic acid I and II and aristolactam I and II using a newly-developed microdialysis liquid chromatography-tandem mass spectrometry.

Aristolochic acids (AAs) are naturally occurring genotoxic carcinogens linked to Balkan endemic nephropathy and aristolochic acid nephropathy. Aristolochic acid I and II (AA-I and AA-II) are the most abundant AAs, and AA-I has been reported to be more genotoxic and nephrotoxic than AA-II. This study aimed to explore metabolic differences underlying the differential toxicity. We developed a novel microdialysis sampling coupled with solid-phase extraction liquid chromatography-tandem mass spectrometry (MD-SPE-LC-MS/MS) to simultaneously study the toxicokinetics (TK) of AA-I and AA-II and their corresponding aristolactams (AL-I and AL-II) in the blood of Sprague Dawley rats co-treated with AA-1 and AA-II. Near real-time monitoring of these analytes in the blood of treated rats revealed that AA-I was absorbed, distributed, and eliminated more rapidly than AA-II. Moreover, the metabolism efficiency of AA-I to AL-I was higher compared to AA-II to AL-II. Only 0.58% of AA-I and 0.084% of AA-II was reduced to AL-I and AL-II, respectively. The findings are consistent with previous studies and support the contention that differences in the in vivo metabolism of AA-I and AA-II may be critical factors for their differential toxicities.

Anti-Cancer Effects of Oxygen-Atom-Modified Derivatives of Wasabi Components on Human Leukemia Cells.

1-Isothiocyanato-6-(methylsulfinyl)-hexanate (6-MITC) is a natural compound found in Wasabia japonica. The synthetic derivatives 1-Isothiocyanato-6-(methylsulfenyl)-hexane (I7447) and 1-Isothiocyanato-6-(methylsulfonyl)-hexane (I7557) were obtained from 6-MITC by deleting and adding an oxygen atom to the sulfone group, respectively. We previously demonstrated that extensive mitotic arrest, spindle multipolarity, and cytoplasmic vacuole accumulation were induced by 6-MITC and inhibited the viability of human chronic myelogenous leukemia K562 cells. In this study, we examined the anti-cancer effects of 6-MITC derivatives on human chronic myelogenous leukemia (CML) cells. Autophagy was identified as the formation of autophagosomes with double-layered membranes using transmission electron microscopy. Cell cycle and differentiation were analyzed using flow cytometry. Apoptosis was detected by annexin V staining. After treatment with I7447 and I7557, the G2/M phase of cell cycle arrest was revealed. Cell death can be induced by a distinct mechanism (the simultaneous occurrence of autophagy and aberrant mitosis). The expression levels of acridine orange were significantly affected by lysosomal inhibitors. The natural wasabi component, 6-MITC, and its synthetic derivatives have similar effects on human chronic myelogenous leukemia cells and may be developed as novel therapeutic agents against leukemia.

Local Liver Irradiation Concurrently Versus Sequentially with Cabozantinib on the Pharmacokinetics and Biodistribution in Rats.

The aim of this study was to evaluate the radiotherapy (RT)-pharmacokinetics (PK) effect of cabozantinib in concurrent or sequential regimens with external beam radiotherapy (EBRT) or stereotactic body radiation therapy (SBRT). Concurrent and sequential regimens involving RT and cabozantinib were designed. The RT-drug interactions of cabozantinib under RT were confirmed in a free-moving rat model. The drugs were separated on an Agilent ZORBAX SB-phenyl column with a mobile phase consisting of 10 mM potassium dihydrogen phosphate (KH2PO4)-methanol solution (27:73, v/v) for cabozantinib. There were no statistically significant differences in the concentration versus time curve of cabozantinib (AUCcabozantinib) between the control group and the RT2Gy×3 f'x and RT9Gy×3 f'x groups in the concurrent and the sequential regimens. However, compared to those in the control group, the Tmax, T1/2 and MRT decreased by 72.8% (p = 0.04), 49.0% (p = 0.04) and 48.5% (p = 0.04) with RT2Gy×3 f'x in the concurrent regimen, respectively. Additionally, the T1/2 and MRT decreased by 58.8% (p = 0.01) and 57.8% (p = 0.01) in the concurrent RT9Gy×3 f'x group when compared with the control group, respectively. The biodistribution of cabozantinib in the heart increased by 271.4% (p = 0.04) and 120.0% (p = 0.04) with RT2Gy×3 f'x in the concurrent and sequential regimens compared to the concurrent regimen, respectively. Additionally, the biodistribution of cabozantinib in the heart increased by 107.1% (p = 0.01) with the RT9Gy×3 f'x sequential regimen. Compared to the RT9Gy×3 f'x concurrent regimen, the RT9Gy×3 f'x sequential regimen increased the biodistribution of cabozantinib in the heart (81.3%, p = 0.02), liver (110.5%, p = 0.02), lung (125%, p = 0.004) and kidneys (87.5%, p = 0.048). No cabozantinib was detected in the brain in any of the groups. The AUC of cabozantinib is not modulated by irradiation and is not affected by treatment strategies. However, the biodistribution of cabozantinib in the heart is modulated by off-target irradiation and SBRT doses simultaneously. The impact of the biodistribution of cabozantinib with RT9Gy×3 f'x is more significant with the sequential regimen than with the concurrent regimen.

Exploration of sodium homeostasis and pharmacokinetics in bile duct-ligated rats treated by anti-cirrhosis herbal formula plus spironolactone.

Renal sodium retention is an essential indicator that is used for the prognosis of cirrhosis with ascites that requires diuretic treatment to restore sodium homeostasis. The diuretic effects of Yin-Chen-Hao-Tang (YCHT) alone or in combination with diuretics for sodium retention in patients with cirrhosis have not been investigated. This study aimed to investigate the diuretic effects and sodium retention caused by YCHT with spironolactone, from both the pharmacokinetic and pharmacodynamic perspective, in bile duct-ligated rats. The HPLC method was validated and utilized for the pharmacokinetic analysis of rat urine. Urine samples were collected and analyzed every 4 hours for 32 h after oral administration of YCHT at 1 or 3 g/kg daily for 5 days in bile duct-ligated rats. A dose of 20 mg/kg spironolactone was also administered to pretreat the YCHT 1 g/kg or the 3 g/kg group on the 5th day to explore the interaction of the two treatments. Urine sodium, potassium, weight, volume, and spironolactone and canrenone levels were measured to investigate fluid homeostasis after the coadministration. The linearity, precision, and accuracy of the HPLC method were suitable for subsequent urinary pharmacokinetic analyses. The pharmacokinetic parameters in the 1 g/kg YCHT with spironolactone group revealed that the elimination half-life of the spironolactone metabolite, canrenone, was prolonged. In addition, the cumulative excretion amount, the area under the rate curve (AURC), and the maximum rate of excretion (Rmax) were significantly decreased when the spironolactone group was pretreated with 3 g/kg YCHT. Urinary sodium excretion elicited by spironolactone was suppressed by pretreatment with 1 or 3 g/kg YCHT. The 32-hour urine output was not altered by the administration of YCHT alone, but it was significantly decreased by 64.9% after the coadministration of YCHT with spironolactone. The interaction of spironolactone and YCHT was found to decrease urine sodium-potassium and water excretion, and this change was attributed to the decreased level of spironolactone metabolites and possibly the regulation of the renin-angiotensin-aldosterone system by obstructed cirrhosis. The dose adjustment of YCHT or diuresis monitoring should be noted when co-administering YCHT and spironolactone to treat hepatic diseases clinically.

Pharmacokinetic and pharmacodynamic herb-drug interactions of common over-the-counter pain medications.

Pain is one of the most common reasons for seeking medical intervention, and self-medication with over-the-counter medications and/or traditional herbal remedies has become increasingly popular. In this review, original articles on understanding possible herb-drug interactions between traditional herbs and four major pain medications-acetaminophen, aspirin, ibuprofen and naproxen-are compiled and analyzed. In terms of analytical methods, high-performance liquid chromatography using an isocratic eluent system coupled to biological sample clean-up is the most common, while a wide variety of detectors have been observed, including a photodiode array, variable wavelength detector, electrochemical detector and tandem mass spectrometer. Both synergistic and anti-synergistic effects were observed for acetaminophen and aspirin, while only synergistic effects have been found for naproxen. Currently, no interactions have been reported for ibuprofen.

Neuroprotective effect and herbal-drug pharmacokinetic interaction of Gastrodia elata extract on valproic acid.

Valproic acid (VPA) is a widely used antiepileptic drug, and the herbal extract of Gastrodia elata exerts an anticonvulsant effect. However, few studies have investigated the pharmacokinetic and pharmacodynamic interactions between G. elata extract and VPA. We hypothesize that G. elata extract increases the VPA levels in the brain and enhances the antiepileptic effects of VPA, and this synergistic effect is mediated by transporters at the bloodbrain barrier (BBB). We performed microdialysis on pilocarpine-induced epileptic model rats in vivo to investigate this hypothesis. The results demonstrated that cotreatment with G. elata extract and VPA ameliorated drug-resistant epilepsy by increasing the VPA levels in the brain. In addition, G. elata extract and VPA exerted synergistic anticonvulsive effects to decrease the seizure severity by protecting neurons in the hippocampus and altering the DOPAC and 5-HT levels. However, these phenomena were partially blocked by the organic anion transporter peptide (OATP) inhibitor cyclosporine A (CsA; 20 mg/kg, i.p.), which demonstrated that the increase in the VPA level in the brain was modulated by the transporter OATP. This study provides a comprehensive strategy for assessing the interaction between traditional medicines and conventional antiepileptic drugs in a status epilepticus animal model.

Transplacental transfer of acetaminophen in pregnant rats.

Acetaminophen is among the most widely used analgesics; however, the proportion and mechanism of transplacental transfer of unbound acetaminophen with actual pharmacological activity remain unknown. Our hypothesis is that acetaminophen gradually penetrates the blood-placenta barrier to reach the fetus. A multiple microdialysis coupled to liquid chromatography with photodiode array detection method was developed to monitor acetaminophen levels in the maternal blood, placenta, fetus, and amniotic fluid of a pregnant rat and investigate this hypothesis. The pharmacokinetic data indicates that acetaminophen exhibits a nonlinear behavior in the maternal blood within the dosage regimen of 100 and 300 mg/kg. In addition, acetaminophen penetrates the placenta, fetus, and amniotic fluid during treatment. The transplacental transfer ratio represented by the area under the concentration curve (AUC) ratio for the conceptus (the collective term for the fetus, placenta, and amniotic fluid) and maternal blood (AUCtissue/AUCblood) was approximately 11-23 % after acetaminophen (100 and 300 mg/kg) administration. However, the transporter of multidrug resistance-associated protein (MRP) inhibitor MK-571 did not significantly change the transplacental transfer ratio. This basic study provides constructive information for the clinical application of acetaminophen in pregnant women.

Wasabi Component 6-(Methylsulfinyl)hexly Isothiocyanate and Derivatives Improve the Survival of Skin Allografts.

We tested the effect of 6-(Methylsulfinyl)hexyl Isothiocyanate (6-MITC) and derivatives (I7447 and I7557) on the differentiation and maturation of human myeloid dendritic cells (DCs) in vitro, and skin transplantation in vivo. Triggering of CD14+ myeloid monocyte development toward myeloid DCs with and without 6-MITC and derivatives to examine the morphology, viability, surface marker expression, and cytokine production. Stimulatory activity on allogeneic naive T cells was measured by proliferation and interferon-γ production. The skin allograft survival area model was used to translate the 6-MITC and derivatives' antirejection effect. All of the compounds had no significant effects on DC viability and reduced the formation of dendrites at concentrations higher than 10 µM. At this concentration, 6-MITC and I7557, but not I7447, inhibited the expression of CD1a and CD83. Both 6-MITC and I7557 exhibited T-cells and interferon-γ augmentation at lower concentrations and suppression at higher concentration. The 6-MITC and I7557 prolonged skin graft survival. Both the 6-MITC and I7557 treatment resulted in the accumulation of regulatory T cells in recipient rat spleens. No toxicity was evident in 6-MITC and I7557 treatment. The 6-MITC and I7557 induced human DC differentiation toward a tolerogenic phenotype and prolonged rat skin allograft survival. These compounds may be effective as immunosuppressants against transplant rejection.

Pharmacokinetics and transplacental transfer of codeine and codeine metabolites from Papaver somniferum L.

ETHNOPHARMACOLOGICAL RELEVANCE: Papaveris Pericarpium, which is the dried husk of Papaver somniferum L., has been used as a phytomedicine to relieve cough, diarrhea and pain. The alkaloid codeine contained therein via biotransformation converts to morphine and potentially produces addictive and toxic effects. Due to the healthy concern for a pregnant woman, our hypothesis is that codeine and its metabolites can penetrate the placental barrier to reach the foetus and amniotic fluid, and these processes may be modulated by the transporter. AIM OF THE STUDY: Because codeine is also considered a prodrug of morphine, it has a good analgesic effect. It is often used by pregnant women but may expose the foetus to the risk of morphine harm. The aim of this study is to investigate the metabolic rate, distribution and transplacental transfer mechanism of codeine and its metabolites morphine and morphine-3-glucuronide (M3G) in pregnant rats and to assess the risk of medication for pregnant women. MATERIALS AND METHODS: Ultrahigh-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) combined with a microdialysis system was developed to monitor codeine, morphine and M3G in multiple sites of maternal blood, placenta, foetus and amniotic fluid after codeine administration. A compartmental model was used to calculate the pharmacokinetic parameters of codeine in blood after codeine administration (10 mg/kg, i.v.). The area under the concentration (AUC) ratio of AUCmetabolite/AUCcodeine and AUCtissue/AUCblood was used to represent the metabolic biotransformation ratio and the drug from blood-to-tissue transfer ratio, respectively. RESULTS: The pharmacokinetic results demonstrated that codeine fit well with a two-compartment model and went through rapid metabolism to morphine and M3G in pregnant rats after codeine administration (10 mg/kg, i.v.). The biotransformation ratios of AUCmorphine/AUCcodeine, AUCM3G/AUCmorphine and AUCM3G/AUCcodeine were 0.12 ± 0.03, 54.45 ± 20.61 and 6.53 ± 2.47, respectively, after codeine administration (10 mg/kg, i.v.), which suggested that codeine was easily metabolized into M3G through morphine. The tissue distribution results demonstrated that all of the analytes penetrated into the foetus through the placenta; however, the blood-to-tissue transfer ratio (AUCtissue/AUCblood) of morphine and M3G was relatively lower than that of codeine after codeine administration (10 mg/kg, i.v.), which suggested that the blood-placenta barrier blocks the penetration of morphine and M3G into the foetus. Thus, the tissue transfer of morphine in the placenta and foetus was significantly enhanced by treatment with corticosterone, an inhibitor of organic cation transporter (OCT). CONCLUSION: Based on microdialysis coupled to a validated UHPLC-MS/MS system, the pharmacokinetics and metabolic biotransformation of codeine and its metabolites were analyzed and clarified. The potential mechanism of morphine placental transfer was modulated by OCT transporters.

Biotransformation and transplacental transfer of the anti-viral remdesivir and predominant metabolite, GS-441524 in pregnant rats.

BACKGROUND: Remdesivir was the first prodrug approved to treat coronavirus disease 2019 (COVID-19) and has the potential to be used during pregnancy. However, it is not known whether remdesivir and its main metabolite, GS-441524 have the potential to cross the blood-placental barrier. We hypothesize that remdesivir and predominant metabolite GS-441524may cross the blood-placental barrier to reach the embryo tissues. METHODS: To test this hypothesis, ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) coupled with multisite microdialysis was used to monitor the levels of remdesivir and the nucleoside analogue GS-441524 in the maternal blood, fetus, placenta, and amniotic fluid of pregnant Sprague-Dawley rats. The transplacental transfer was evaluated using the pharmacokinetic parameters of AUC and mother-to-fetus transfer ratio (AUCfetus/AUCmother). FINDINGS: Our in-vivo results show that remdesivir is rapidly biotransformed into GS-441524 in the maternal blood, which then readily crossed the placenta with a mother-to-fetus transfer ratio of 0.51 ± 0.18. The Cmax and AUClast values of GS-441524 followed the order: maternal blood > amniotic fluid > fetus > placenta in rats. INTERPRETATION: While remdesivir does not directly cross into the fetus, however, its main metabolite, GS-441524 readily crosses the placenta and can reside there for at least 4 hours as shown in the pregnant Sprague-Dawley rat model. These findings suggest that careful consideration should be taken for the use of remdesivir in the treatment of COVID-19 in pregnancy. FUNDING: Ministry of Science and Technology of Taiwan.

Impact of Irradiation on the Pharmacokinetics and Biotransformation of Tamoxifen.

BACKGROUND: The optimal procedure for combining radiotherapy (RT) with tamoxifen treatment is controversial as RT may alter the pharmacokinetics and biotransformation of tamoxifen. The present study investigated this potential interaction by assessing the pharmacokinetics of tamoxifen during concurrent and sequential RT. METHOD: Plasma tamoxifen concentration was measured in rats with or without RT 2.0 Gy (RT2.0Gy) or 0.5 Gy (RT0.5Gy) with ultra-high-performance liquid chromatography-tandem mass spectrometry after tamoxifen administration (10 mg/kg, p.o., n = 6). Tamoxifen was either administered 1 h after RT (concurrent condition) or 24 h after RT (sequential condition). RESULTS: Pharmacokinetic data analysis demonstrated that the area under the curve (AUC) and half-life of tamoxifen were 2,004 ± 241 h ng/ml and 6.23 ± 1.21 h, respectively, after tamoxifen administration (10 mg/kg, p.o.). The respective conversion rate of 4-hydroxytamoxifen, N-desmethytamoxifen, and endoxifen for tamoxifen metabolism was 20%, 16%, and 5%. The AUC value of tamoxifen in the RT0.5Gy group was 1.5- to 1.7-fold higher than in the sham and RT2.0Gy groups. The relative bioavailability of tamoxifen at concurrent RT0.5Gy and RT2.0Gy groups ranged from 127% to 202% and from 71% to 152%, respectively. The magnitude of endoxifen, which converted from 4-hydroxytamoxifen and N-desmethyltamoxifen, increased 3- to 5-fold in the concurrent RT groups. By contrast, the AUC of tamoxifen decreased by roughly 24% in the sequential RT2.0Gy group. The conversion ratio of endoxifen was four times higher than that in the sequential RT2.0Gy group compared with rats not exposed to RT. CONCLUSION: The current study provides advanced pharmacokinetic data to confirm the interaction between RT and hormone therapy. Our findings indicate that RT facilitates the metabolism of tamoxifen to active metabolites and thus imply that combination RT-tamoxifen has potential benefits for the treatment of hormone-dependent breast cancer.

Borneol and Tetrandrine Modulate the Blood-Brain Barrier and Blood-Tumor Barrier to Improve the Therapeutic Efficacy of 5-Fluorouracil in Brain Metastasis.

The efficacy of chemotherapeutic drugs for the treatment of brain metastasis may be compromised by the blood-brain barrier (BBB) and blood-tumor barrier (BTB). P-glycoprotein (P-gp) is a multidrug resistance protein that potentially limits the penetration of chemotherapeutics through the BBB and BTB. 5-Fluorouracil (5-FU) is widely used to treat cancer. Bioactive constituents of medicinal herbs, such as borneol and tetrandrine, potentially improve drug penetration through the BBB and BTB. We hypothesized that borneol and tetrandrine might modulate the BBB and BTB to enhance 5-FU penetration into the brain. To investigate this, in vitro and in vivo models were developed to explore the modulatory effects of borneol and tetrandrine on 5-FU penetration through the BBB and BTB. In the in vitro models, barrier integrity, cell viability, barrier penetration, P-gp activity, and NF-κB expression were assessed. In the in vivo brain metastasis models, cancer cells were injected into the internal carotid artery to evaluate tumor growth. The experimental results demonstrated that borneol and borneol + tetrandrine reduced BBB integrity. The efflux pump function of P-gp was partially inhibited by tetrandrine and borneol + tetrandrine. In the in vivo experiment, borneol + tetrandrine effectively prolonged survival without compromising body weight. In conclusion, BBB and BTB integrity was modulated by borneol and borneol + tetrandrine. The combination of borneol and tetrandrine could be used to improve the chemotherapeutic control of brain metastasis.

Metformin-Incorporated Gelatin/Hydroxyapatite Nanofiber Scaffold for Bone Regeneration.

Tissue engineering and regenerative medicine has gradually evolved as a promising therapeutic strategy to the modern health care of aging and diseased population. In this study, we developed a novel nanofibrous scaffold and verified its application in the critical bone defect regeneration. The metformin-incorporated nano-gelatin/hydroxyapatite fibers (NGF) was produced by electrospinning, cross-linked, and then characterized by X-ray powder diffractometer and Fourier-transform infrared spectroscopy. Cytotoxicity, cell adhesion, cell differentiation, and quantitative osteogenic gene and protein expression were analyzed by bone marrow stem cells (BMSCs) from rat. Rat forearm critical bone defect model was performed for the in vivo study. The NGF were characterized by their porous structures with proper interconnectivity without significant cytotoxic effects; the adhesion of BMSCs on the NGF could be enhanced. The osteogenic gene and protein expression were upregulated. Postimplantation, the new regenerated bone in bone defect was well demonstrated in the NGF samples. We demonstrated that the metformin-incorporated NGF greatly improved healing potential on the critical-size bone defect. Although metformin-incorporated NGF had advantageous effectiveness during bone regeneration, further validation is required before it can be applied to clinical applications. Impact statement Bone is the structure that supports the rest of the human body. Critical-size bone defect hinders the regeneration of damaged bone tissues and compromises the mechanical strength of the skeletal system. Characterized by their porous structures with proper interconnectivity, the electrospinning nano-gelatin/hydroxyapatite fibrous scaffold developed in this study can greatly improve the healing potential on the critical-size bone defect. Further validation can validate its potential clinical applications.

Simultaneous toxicokinetics characterization of acrylamide and its primary metabolites using a novel microdialysis isotope-dilution liquid chromatography mass spectrometry method.

Acrylamide (AA) is a toxicant in high-temperature processed foods and an animal carcinogen. Upon absorption, AA is metabolized to glycidamide (GA) or conjugates with glutathione (AA-GSH). Important advantages of microdialysis coupled with liquid chromatography-tandem mass spectrometry (MD-LC-MS/MS) include its minimization of potential losses during sample collection, storage and preparation, as well as an improvement in temporal resolution for toxicokinetics (TKs). We aimed to simultaneously study the TKs of AA and products of its primary metabolism using an isotope-dilution (ID) MD-LC-MS/MS method. MD probes implanted into the jugular vein/right atrium of anesthetized Sprague Dawley rats were connected to the ID-LC-MS/MS for continuous monitoring of AA, GA and AA-GSH in the blood every 15 min over 8 h following intraperitoneal AA administration (0.1 mg/kg or 5 mg/kg). AA, GA, and AA-GSH TKs followed linear kinetics: GA AUC/AA AUC = 0.11 and AA-GSH AUC/AA AUC = 0.011 at 5 mg/kg. Elimination half-life (Te1/2) values were 2.44 ± 0.70, 4.93 ± 2.37 and 3.47 ± 1.47 h for AA, GA and AA-GSH, respectively. GA TKs reached a plateau at 3-6 h, suggesting that metabolic saturation of AA and Te1/2 values of the analytes were prolonged with AA at 5 mg/kg. Our results demonstrate that oxidation of AA to GA overwhelmed the conjugation of AA with GSH. Our innovative MD-ID-LC-MS/MS method facilitates the simultaneous characterization of multiple TKs associated with toxicants and their active metabolites with excellent temporal resolution to capture metabolic saturation of AA to GA.