Paclitaxel-Containing Extract Exerts Anti-Cancer Activity through Oral Administration in A549-Xenografted BALB/C Nude Mice: Synergistic Effect between Paclitaxel and Flavonoids or Lignoids.

Taxus yunnanensis is a paclitaxel-containing herb with traditional usage in cancer treatment, and its extract possesses great oral bioavailability of paclitaxel. However, it is elusive whether paclitaxel-containing extract (HDS-1) can exert anti-tumor effect through oral administration and how other components contribute to its efficacy. Therefore, we investigate the oral-route anti-tumor effect of HDS-1 in A549-bearing mice. HDS-1-derived flavonoids (HDS-2) and lignoids (HDS-3) are hypothesized to contribute to HDS-1's efficacy, and their effects of enhancing enterocytic absorption and cytotoxicity of paclitaxel are validated in 2 permeability experiments and apoptosis-related assay, respectively. In vivo, A549 growth is significantly inhibited by 86.1 ± 12.94% (P < 0.01) at 600 mg/kg of HDS-1 and 65.7 ± 38.71% (P < 0.01) at 200 mg/kg. HDS-2 and HDS-3 significantly reduce the efflux ratio of paclitaxel to 2.33 and 3.70, respectively, in Caco-2 permeability experiment and reduce paclitaxel reflux in MDCK-MDR1 experiment. Furthermore, HDS-2 and HDS-3 potentiated paclitaxel-induced cytotoxicity by 19.1-22.45% (P < 0.05) and 10.52-18.03% (P < 0.05), respectively, inhibited the expression of cyclinB1, Bcl-2, and pMCL-1, and increased the percentage of necrosis cell in the condition of paclitaxel exposure. Conclusively, paclitaxel-containing extracts exert anti-cancer effects through oral administration, and flavonoid and lignoids contribute to its anti-cancer effect through simultaneously improving enterocytic absorption of paclitaxel and the cytotoxic effect of paclitaxel.

Simultaneous determination of curcumin, tetrahydrocurcumin, quercetin, and paeoniflorin by UHPLC-MS/MS in rat plasma and its application to a pharmacokinetic study.

Curcumin (CUR) is a bioactive compound present in many composite prescriptions of traditional Chinese medicine together with quercetin (QR) and paeoniflorin (PF). Little is known about the influence of QR and PF on the absorption and metabolism of CUR when the three compounds are orally co-administered. In this study, a rapid, sensitive, and reliable ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for the simultaneous determination of CUR, tetrahydrocurcumin (THC), QR, and PF in rat plasma by using tinidazole as the internal standard (IS). A liquid-liquid extraction method with ethyl acetate was used to pre-treat the plasma samples. Chromatographic separation was conducted on a C18 column with isocratic elution using acetonitrile and 0.1% formic acid water solution (80:20, v/v) as the mobile phase at the flow rate of 0.3 mL/min. A TSQ Quantum Access Max API mass spectrometer equipped with electrospray ionisation (ESI) source in selection reaction monitoring (SRM) mode was employed to determine transitions of m/z 369.0 → 176.9, 373.1 → 137.0, 303.0 → 228.9, 478.9 → 120.9, 248.1 → 121.0 for CUR, THC, QR, PF, and IS, respectively. The selectivity, precision, accuracy, extraction recovery, matrix effect, and stability of the method were validated. This developed and validated method was successfully applied in the pharmacokinetic study of CUR, THC, QR, and PF in rats. The effects of QR and PF on the pharmacokinetics of CUR and its metabolite, THC, were evaluated in the plasma of Sprague-Dawley rats that were orally co-administered CUR, QR, and PF. The results showed that the combined use of QR, PF, and CUR has a possible influence on the metabolism and excretion of CUR. Our work provides a fundamental method for the rapid simultaneous determination of CUR, THC, QR, and PF in rat plasma. Furthermore, this study will provide a basic method for the analysis of pharmacokinetic interaction of CUR, QR, and PF and offer a scientific basis for a possible combination therapy with the three compounds.

Mulberroside A suppresses PXR-mediated transactivation and gene expression of P-gp in LS174T cells.

Mulberroside A (Mul A) is the main bioactive constituents of Sangbaipi, which is officially listed in the Chinese Pharmacopoeia. The pregnane X receptor (PXR) has been recognized as the critical mediator of human P-glycoprotein (P-gp) gene transactivation. In this study, the effect of Mul A on PXR-mediated transactivation and gene expression of P-gp was investigated. It was found that Mul A significantly suppressed PXR-mediated P-gp luciferase activity induced by rifampicin (Rif). Furthermore, Rif induced an elevation of P-gp expression and transport activity, which was apparently suppressed by Mul A. However, Mul A did not suppress the P-gp luciferase activity, P-gp expression, and function in the absence of Rif. These findings suggest that Mul A suppresses PXR-mediated transactivation and P-gp expression induced by Rif. This should be taken into consideration to predict any potential herb-drug interactions when Mul A or Sangbaipi are co-administered with Rif or other PXR agonist drugs.

Mechanistic understanding of the effect of Dengzhan Shengmai capsule on the pharmacokinetics of clopidogrel in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: The Dengzhan Shengmai capsule (DZSM) is known in China for its remarkable curative effect as a treatment of cardiovascular diseases, such as coronary heart disease and ischemic stroke. DZSM is a Chinese herbal compound preparation that consists of four ingredients, including Erigeron breviscapus (Vaniot) Hand.-Mazz., Panax ginseng C.A. Mey, Ophiopogon japonicas (Thunb.) Ker-Gawl. and Schisandra chinensis (Turcz.) Baill., and was indexed in the Chinese Pharmacopoeia 2010. DZSM and clopidogrel are often co-prescribed in the clinic to prevent the recurrence of stroke or other cardiovascular and cerebrovascular diseases. However, the effect of DZSM on the pharmacokinetics of clopidogrel remains unclear. AIM OF THE STUDY: The purpose of the study is to explore the pharmacokinetics and potential interaction between DZSM and clopidogrel and the underlying mechanism. MATERIALS AND METHODS: Rats were used to investigate the effect of DZSM on the pharmacokinetics of clopidogrel and its active metabolite in vivo. The plasma concentrations were simultaneously determined using LC-MS/MS. The effects of DZSM on the P-gp-mediated efflux transport and CYP450-mediated metabolism of clopidogrel were investigated using MDCKII-MDR1 cells and rat liver microsomes, respectively. RESULTS: After pretreatment with DZSM, the Cmax and AUC0-∞ of clopidogrel increased from 0.4±0.1 to 1.7±0.6ng/mL and 0.9±0.4 to 2.0±0.2ng/mLh, respectively. The Cmax and AUC0-∞ of the derivatized active metabolite of clopidogrel decreased from 8.2±1.2 to 2.8±0.5ng/mL and 18.2±5.6 to 6.4±3.7ngh/mL, respectively. In MDCKII-MDR1 cells, the P-gp-mediated efflux transport of clopidogrel was significantly inhibited by the DZSM extract. In rat liver microsomes, DZSM inhibited clopidogrel metabolism with an IC50 of 0.02mg/mL. CONCLUSIONS: DZSM significantly affects the pharmacokinetics of clopidogrel and its active metabolite by inhibiting the P-gp-mediated efflux transport and CYP450-mediated metabolism of clopidogrel. Thus, caution is needed when DZSM is co-administered with clopidogrel in the clinic because the interaction of these drugs may result in altered plasma concentrations of clopidogrel and its active metabolite.

Simultaneous determination of seven taxoids in rat plasma by UPLC-MS/MS and pharmacokinetic study after oral administration of Taxus yunnanensis extracts.

A rapid, sensitive and reliable method has been developed and validated for the simultaneous determination of seven taxoids including 10-deacetylbaccatin III (10-DAB III), baccatin III, 5-epi-canadensene, taxinine M, 10-deacetyltaxol (10-DAT), cephalomannine and paclitaxel in rat plasma using docetaxel as the internal standard (IS). The plasma samples were pretreated by liquid-liquid extraction with methyl tert-butyl ether. The chromatographic separation was achieved on a C18 column (50 mm × 2.1 mm, 1.8 µm, Waters, USA) with a gradient elution program consisting of methanol and water (containing 0.1% formic acid) at a flow rate of 0.2 mL/min. Detection was performed under the selected reaction monitoring (SRM) scan using an electrospray ionization (ESI) in the positive ion mode. The mass transitions were as follows: m/z 567.4→444.9 for 10-DAB III, m/z 609.0→549.3 for baccatin III, m/z 617.4→496.9 for 5-epi-canadensene, m/z 709.6→649.3 for taxinine M, m/z 834.8→307.9 for 10-DAT, m/z 854.5→285.4 for cephalomannine, m/z 876.8→307.3 for paclitaxel and m/z 830.8→549.6 for IS, respectively. All calibration curves exhibited good linearity (r(2)>0.99) over a wide concentration range for all components. The intra-day and inter-day precisions at three different levels were both less than 14.3% in terms of relative standard deviation (RSD) and the accuracies ranged from -8.3% to 14.8% in terms of relative error (RE). The extraction recoveries of the seven compounds ranged from 62.5% to 100.5%. The developed method was successfully applied to the pharmacokinetic study of the seven taxoids in rat plasma after oral administration of the crude extract of the twigs and leaves of Taxus yunnanensis.

Effect of scutellarin on the metabolism and pharmacokinetics of clopidogrel in rats.

Erigeron breviscapus (Vant.) Hand-Mazz, a traditional Chinese medicine, is often co-prescribed with clopidogrel for the treatment of ischemic vascular diseases. Scutellarin is the representative bioactive flavonoid isolated from this herb. The aim of this study was to explore the effect of scutellarin on the metabolism and pharmacokinetics of clopidogrel. The in vitro studies using rat liver microsomes showed that scutellarin significantly inhibited the metabolism of clopidogrel. The IC50 value was 2.1 µM. Ten male rats were employed to investigate the effect of scutellarin on the pharmacokinetics of clopidogrel in vivo. After pretreatment with scutellarin, there were significant increases in the AUC0-∞ (from 0.9 ± 0.4 to 1.7 ± 0.6 ng/ml h; p <0.05) and Cmax (from 0.4 ± 0.1 to 0.9 ± 0.1 ng/ml; p <0.05) of clopidogrel. The pharmacokinetic data for clopidogrel active metabolite showed significant decreases in AUC0-∞ (18.2 ± 5.6 to 11.4 ± 3.7 ng/ml h; p <0.05) and Cmax (from 8.2 ± 1.2 to 4.3 ± 0.3 ng/ml; p <0.05) after pretreatment with scutellarin. Collectively, the metabolism and pharmacokinetics of clopidogrel were significantly affected by scutellarin. This study indicated that potential herb-drug interaction between scutellarin and clopidogrel should be taken into consideration in clinical use.

Down-regulation of P-gp expression and function after Mulberroside A treatment: potential role of protein kinase C and NF-kappa B.

P-Glycoprotein (P-gp) plays a major role in drug-drug and herb-drug interactions. Mulberroside A (Mul A) is one of the main bioactive constituents of Sangbaipi, the dried root-bark of Morus alba L. (white mulberry), which is officially listed in the Chinese Pharmacopoeia. In the present study, we investigated the effect of Mul A treatment on mRNA expression and protein expression of P-gp in the Caco-2 cells by real-time qPCR and Western blot analysis. The effect of Mul A treatment on the function of P-gp in vitro and in vivo was assessed by Rho123 transport assay and a pharmacokinetic study. The potential roles of protein kinase C (PKC) and nuclear factor kappa B (NF-κB) in the expression regulation of P-gp after Mul A treatment were also investigated. The results revealed that Mul A treatment significantly decreased the mRNA and protein expression of P-gp in Caco-2 cells after treatment with Mul A (5-20 µM). Furthermore, Mul A treatment displayed apparently inhibitory effect on the function of P-gp both in vitro and in vivo. In addition, activation of PKC activity and NF-κB nuclear translocation were observed in the presence of Mul A, which suggested that PKC and NF-κB might play crucial roles in Mul A-induced suppression of P-gp. Our study demonstrated that Mul A treatment could down-regulate P-gp expression and function accompanied by the activation of PKC and NF-κB, and this should be taken into consideration in potential herb-drug interactions when Mul A or M. alba are co-administered with other drugs transported by P-gp.

Effect of Tacrolimus on the pharmacokinetics of bioactive lignans of Wuzhi tablet (Schisandra sphenanthera extract) and the potential roles of CYP3A and P-gp.

We recently reported that Wuzhi tablet (WZ), a preparation of the ethanol extract of Wuweizi (Schisandra sphenanthera), had significant effects on blood concentrations of Tacrolimus (FK506) in renal transplant recipients and rats. The active lignans in WZ are schisandrin A, schisandrin B, schisandrin C, schisandrol A, schisandrol B, schisantherin A, and schisantherin B. Until now, whether the pharmacokinetics of these lignans in WZ would be affected by FK506 remained unknown. Therefore, this study aimed to investigate whether and how FK506 affected pharmacokinetics of lignans in WZ in rats and the potential roles of CYP3A and P-gp. After a single oral co-administration of FK506 and WZ, the blood concentration of lignans in WZ was decreased by FK506; furthermore, the AUC of schisantherin A, schisandrin A, schisandrol A and schisandrol B was only 64.5%, 47.2%, 55.1% and 57.4% of that of WZ alone group, respectively. Transport study in Caco-2 cells showed that these lignans were not substrates of P-gp, suggesting decreased blood concentration of lignans by FK506 was not via P-gp pathway. Metabolism study in the human recombinant CYP 3A showed that these lignans had higher affinity to CYP3A than that of FK506, and thus had a stronger CYP3A-mediated metabolism. It was concluded that the blood concentrations of these lignans were decreased and their CYP3A-mediated metabolisms were increased in the presence of FK506 since these lignans had higher affinity to CYP3A.

Comparative pharmacokinetics of paclitaxel after oral administration of Taxus yunnanensis extract and pure paclitaxel to rats.

Taxus yunnanensis (T. yunnanensis) is endemic to China and has been used in traditional medicine for the treatment of cancer, diabetic ailments and others. Paclitaxel is a representative antitumor compound in the Taxus species. The pharmacokinetic behavior of paclitaxel after oral administration of the crude extract of T. yunnanensis has not been investigated. This study attempts to compare the pharmacokinetics of paclitaxel after an oral administration of the crude extract of the twigs and leaves of T. yunnanensis and pure paclitaxel. A UPLC and a UPLC/MS/MS analysis method were developed for the determination of paclitaxel in T. yunnanensis extract and in the comparative pharmacokinetic study. Caco-2 cells were used to investigate the transport profile of paclitaxel in vitro. In the pharmacokinetic study, rats were randomly grouped and administered with T. yunnanensis extract or pure paclitaxel. The results showed that the AUC and C(max) of paclitaxel in rats receiving the T. yunnanensis extract were significantly increased than those receiving the pure paclitaxel, and the in vitro Caco-2 cell monolayer transport study found that the coexisting constituents in the extract of T. yunnanensis could inhibit the efflux of paclitaxel. These findings suggested that the oral absorption and bioavailability of paclitaxel in T. yunnanensis extract were remarkably higher when compared with the pure paclitaxel, and the coexisting constituents in the T. yunnanensis extract might play an important role for the enhancement of the oral absorption and bioavailability of paclitaxel.

Effect of Wuzhi tablet (Schisandra sphenanthera extract) on the pharmacokinetics of cyclosporin A in rats.

In our previous reports, Wuzhi tablet (an herbal preparation of ethanol extract of Wuweizi (Schisandra sphenanthera)) can significantly increase the blood concentration of tacrolimus and paclitaxel in rats by inhibiting the CYP3A-mediated metabolism and the P-gp-mediated efflux. Cyclosporin A (CsA), a well-known immunosuppressant agent, is also a substrate of CYP3A and P-gp. Therefore, this study aimed to investigate whether and how WZ affects pharmacokinetics of CsA in rats. The AUC0-48 h and Cmax of CsA were increased by 40.1% and 13.1%, respectively, with a single oral co-administration of WZ and high dose of CsA (37.8 mg/kg). Interestingly, after a single oral co-administration of WZ and low dose of CsA (1.89 mg/kg), the AUC0-36 h and Cmax of CsA were dramatically increased by 293.1% (from 1103.2 ± 293.0 to 4336.5 ± 1728.3 ng.h/mL; p < 0.05) and 84.1% (from 208.5 ± 67.9 to 383.1 ± 92.5 ng/mL; p < 0.05), respectively. The CL/F was decreased from 1.7 L/h/kg to 0.5 L/h/kg. Thus, the effect of WZ on high dose of CsA was not significant, but pharmacokinetic parameters of CsA at low dose were significantly influenced by co-administration of WZ. The herb-drug interaction should be taken into consideration at this situation.

Effect of Wuzhi tablet (Schisandra sphenanthera extract) on the pharmacokinetics of paclitaxel in rats.

Wuzhi tablet (WZ, registration no. in China: Z20025766) is a preparation of an ethanol herb extract of Wuweizi (Schisandra sphenanthera) containing 7.5 mg Schisantherin A per tablet. It was reported recently that WZ could significantly increase the blood concentrations of tacrolimus, which might be due to the inhibitory effect of WZ and its ingredients on P-gp and/or CYP450 activity. Paclitaxel is a substrate of the efflux transporter P-gp, and is mainly metabolized by CYP450 enzymes in the liver. Therefore, the purpose of this study was to investigate whether and how WZ affects the pharmacokinetics of paclitaxel in rats. After pretreatment with WZ, there were significant increases in the AUC(0-24h) of oral paclitaxel (from 280.8 ± 97.3 to 543.5 ± 115.2 h ng/mL; p < 0.05) and C(max) (from 44.6 ± 16.4 to 86.8 ± 16.1 ng/mL; p < 0.05). The pharmacokinetic data for i.v. paclitaxel with WZ showed a relatively small (when compared against oral paclitaxel) but still significant increase in AUC(0-24h) (from 163.6 ± 22.1 to 212.7 ± 17.7 h ng/mL; p < 0.05) and a decrease in clearance (from 3.2 ± 0.6 to 2.2 ± 0.3 L/h/kg; p < 0.05). Thus, the presence of WZ improved the systemic exposure of paclitaxel in rats. The herb-drug interaction between WZ and paclitaxel should be taken into consideration in clinical use.