Electroacupuncture stimulation enhances the permeability of the blood-brain barrier: A systematic review and meta-analysis of preclinical evidence and possible mechanisms.

An important cellular barrier to maintain the stability of the brain's internal and external environment is the blood-brain barrier (BBB). It also prevents harmful substances from entering brain tissue through blood circulation while providing protection for the central nervous system. It should be noted, however, that the intact BBB can be a barrier to the transport of most drugs into the brain via the conventional route of administration, which can prevent them from reaching effective concentrations for the treatment of disorders affecting the central nervous system. Electroacupuncture stimulation has been shown to be effective at opening the BBB in a series of experimental studies. This study systematically analyzes the possibility and mechanism by which electroacupuncture opens the BBB. In PubMed, Web of Science, VIP Database, Wanfang Database, and the Chinese National Knowledge Infrastructure, papers have been published for nearly 22 years aimed at opening the BBB and its associated structures. A comparison of EB content between electroacupuncture and control was selected as the primary outcome. There were also results on vascular endothelial growth factor (VEGF), nerve growth factor (NGF), P-Glycoprotein (P-gp), Matrix Metalloproteinase 9 (MMP-9), and glial fibrillary acidic protein (GFAP). We utilized Review Manager software analysis to analyze correlations between studies with a view to exploring the mechanisms of similarity. Evans Blue infiltration forest plot: pooled effect size of 2.04, 95% CI: 1.21 to 2.87, P < 0.01. These results indicate that electroacupuncture significantly increases EB penetration across the BBB. Most studies have reported that GFAP, MMP-9, and VEGF were upregulated after treatment. P-gp expression decreased as well. Electroacupuncture can open the BBB, and the sparse-dense wave is currently the most effective electroacupuncture frequency for opening the BBB. VEGF plays an important role in opening the BBB. It is also important to regulate the expression of MMP-9 and GFAP and inhibit the expression of P-gp.

Lycorine and homolycorine derivatives for chemo-sensitizing resistant human ovarian adenocarcinoma cells.

BACKGROUND: Multidrug resistance is the major obstacle to cancer chemotherapy. Modulation of P-glycoprotein and drug combination approaches have been considered important strategies to overcome drug resistance. PURPOSE: Aiming at generating a small library of Amaryllidaceae-type alkaloids to overcome drug resistance, two major alkaloids, isolated from Pancratium maritimum, lycorine (1), and 2α-10bα-dihydroxy-9-O-demethylhomolycorine (2), were derivatized, giving rise to nineteen derivatives (3 - 21). METHODS: The main chemical transformation of lycorine resulted from the cleavage of ring E of the diacetylated lycorine derivative (3) to obtain compounds that have carbamate and amine functions (5 - 16), while acylation of compound 2 provided derivatives 17 - 21. Compounds 1 - 21 were evaluated for their effects on cytotoxicity, and drug resistance reversal, using resistant human ovarian carcinoma cells (HOC/ADR), overexpressing P-glycoprotein (P-gp/ABCB1), as model. RESULTS: Excluding lycorine (1) (IC50 values of 1.2- 2.5 µM), the compounds were not cytotoxic or showed moderate/weak cytotoxicity. Chemo-sensitization assays were performed by studying the in vitro interaction between the compounds and the anticancer drug doxorubicin. Most of the compounds have shown synergistic interactions with doxorubicin. Compounds 5, 6, 9 - 14, bearing both carbamate and aromatic amine moieties, were found to have the highest sensitization rate, reducing the dose of doxorubicin 5-35 times, highlighting their potential to reverse drug resistance in combination chemotherapy. Selected compounds (4 - 6, 9 - 14, and 21), able of re-sensitizing resistant cancer cells, were further evaluated as P-gp inhibitors. Compound 11, which has a para­methoxy-N-methylbenzylamine moiety, was the strongest inhibitor. In the ATPase assay, compounds 9-11 and 13 behaved as verapamil, suggesting competitive inhibition of P-gp. At the same time, none of these compounds affected P-gp expression at the mRNA or protein level. CONCLUSIONS: This study provided evidence of the potential of Amaryllidaceae alkaloids as lead candidates for the development of MDR reversal agents.

Targeted inhibition of the HNF1A/SHH axis by triptolide overcomes paclitaxel resistance in non-small cell lung cancer.

Paclitaxel resistance is associated with a poor prognosis in non-small cell lung cancer (NSCLC) patients, and currently, there is no promising drug for paclitaxel resistance. In this study, we investigated the molecular mechanisms underlying the chemoresistance in human NSCLC-derived cell lines. We constructed paclitaxel-resistant NSCLC cell lines (A549/PR and H460/PR) by long-term exposure to paclitaxel. We found that triptolide, a diterpenoid epoxide isolated from the Chinese medicinal herb Tripterygium wilfordii Hook F, effectively enhanced the sensitivity of paclitaxel-resistant cells to paclitaxel by reducing ABCB1 expression in vivo and in vitro. Through high-throughput sequencing, we identified the SHH-initiated Hedgehog signaling pathway playing an important role in this process. We demonstrated that triptolide directly bound to HNF1A, one of the transcription factors of SHH, and inhibited HNF1A/SHH expression, ensuing in attenuation of Hedgehog signaling. In NSCLC tumor tissue microarrays and cancer network databases, we found a positive correlation between HNF1A and SHH expression. Our results illuminate a novel molecular mechanism through which triptolide targets and inhibits HNF1A, thereby impeding the activation of the Hedgehog signaling pathway and reducing the expression of ABCB1. This study suggests the potential clinical application of triptolide and provides promising prospects in targeting the HNF1A/SHH pathway as a therapeutic strategy for NSCLC patients with paclitaxel resistance. Schematic diagram showing that triptolide overcomes paclitaxel resistance by mediating inhibition of the HNF1A/SHH/ABCB1 axis.

Mosloflavone from Fissistigma petelotii ameliorates oncogenic multidrug resistance by STAT3 signaling modulation and P-glycoprotein blockade.

BACKGROUND: Oncogenic multidrug resistance (MDR) is a tough question in cancer therapy. However, no effective medications targeting oncogenic MDR are currently available. Studies have demonstrated that mosloflavone exerts anti-inflammatory effects, yet, its potential to ameliorate MDR remains unclear. PURPOSE: This study aimed to access the capability and elucidate molecular mechanisms of mosloflavone as a MDR resensitizing candidate. METHODS: We investigated the ability of mosloflavone to reverse oncogenic MDR and investigated its underlying mechanisms through cytotoxicity assay, cell cycle assay, apoptosis assay, and zebrafish xenograft model. The modulatory interplay between mosloflavone and P-gp was investigated through analysis of calcein-AM uptake, substrate efflux, ATPase assays, and molecular docking simulation. RESULTS: Mosloflavone inhibited P-gp efflux function in an uncompetitive manner without altering ABCB1 gene expression. In addition, it stimulated P-gp ATPase activity by binding to an active site distinct from that of verapamil. Regarding MDR reversal potential, mosloflavone resensitized MDR cancer cells to chemotherapies by arresting cell cycle and triggering apoptosis, possibly by enhancing reactive oxygen species accumulation and reducing phospho-STAT3. Moreover, in the zebrafish xenograft model, mosloflavone significantly potentiated the antitumor effect of paclitaxel. CONCLUSION: Our findings underscore the potential of mosloflavone as a novel dual modulator of STAT3 and P-gp, indicating it is a promising candidate for overcoming MDR in cancer treatment.

Overcoming multi drug resistance mediated by ABC transporters by a novel acetogenin- annonacin from Annona muricata L.

ETHNOPHARMACOLOGICAL RELEVANCE: Multi-Drug Resistance (MDR), mediated by P-glycoprotein (P-gp) is one of the barriers to successful chemotherapy in colon cancer patients. Annona muricata L. (A.muricata), commonly known as soursop/Graviola, is a medicinal plant that has been traditionally used in treating diverse diseases including cancer. Phytochemicals of A.muricata (Annonaceous Acetogenins-AGEs) have been well-reported for their anti-cancer effects on various cancers. AIM OF THE STUDY: The study aimed to examine the effect of AGEs in reversing MDR in colorectal cancer cells. METHODS: Based on molecular docking and molecular dynamic simulation, the stability of annonacin upon P-gp was investigated. Further in vitro studies were carried in oxaliplatin-resistant human colon cancer cells (SW480R) to study the biological effect of annonacin, in reversing drug resistance in these cells. RESULTS: Molecular docking and simulation studies have indicated that annonacin stably interacted at the drug binding site of P-gp. In vitro analysis showed that annonacin was able to significantly reduce the expression of P-gp by 2.56 folds. It also induced apoptosis in the drug-resistant colon cancer cells. Moreover, the intracellular accumulation of P-gp substrate (calcein-AM) was observed to increase in resistant cells upon treatment with annonacin. CONCLUSION: Our findings suggest that annonacin could inhibit the efflux of chemotherapeutic drugs mediated by P-gp and thereby help in reversing MDR in colon cancer cells. Further in vivo studies are required to decipher the underlying mechanism of annonacin in treating MDR cancers.

Vitamin D3 supplementation may attenuate morphological and molecular abnormalities of the olfactory bulb in a mouse model of Down syndrome.

Individuals with Down syndrome (DS) exhibit impaired olfactory function and are at a higher risk of developing Alzheimer's disease (AD). Olfactory dysfunction may be an early clinical symptom of AD. Recent studies have demonstrated that vitamin D3 (VD3) exerts neuroprotective effects in mouse models of AD. In this study, we investigated the effects of VD3 on the morphology, immunolocalization, and markers involved in neuropathogenic processes, apoptosis, proliferation, cell survival, and clearance of amyloid peptides, along with neuronal markers in the olfactory bulb (OB) of an adult female mouse model of DS. Morphological and molecular analyses revealed that trisomic mice exhibited a volume reduction in the external plexiform layer, a decrease in the number of mitral and granule cells, and an increase in the expression of amyloid-ß 42, caspase-3 p12, and P-glycoprotein. VD3 reversed certain morphological abnormalities in the OB of control trisomic mice (Ts(CO)) and decreased the levels of caspase-3 p12 and methylenetetrahydrofolate reductase in the treated groups. The results demonstrated that trisomy factor causes morphofunctional abnormalities in the OB of Ts(CO) mice. Moreover, VD3 could represent a therapeutic target to attenuate morphological and molecular alterations in OB.

Mahanimbine isolated from Murraya koenigii inhibits P-glycoprotein involved in lung cancer chemoresistance.

P-glycoprotein (P-gp), a transmembrane glycoprotein, is mainly involved in lung cancer multidrug resistance. Several P-gp inhibitors have been developed to enhance the efficacy of chemotherapeutics and overcome drug resistance. However, most of them failed in the clinical stages due to undesirable side effects. Therefore, there is a requirement to develop P-gp inhibitors from natural sources. Dietary spice bioactives have been well-known for their anticancer activities. However, their role in modulating the P-gp activity has not been well investigated. Therefore, we have screened for the potential bioactives from various spice plants with P-gp modulatory activity using computational molecular docking analysis. The computational analysis revealed several key bioactives from curry leaves, specifically mahanimbine, exhibited a strong binding affinity with P-gp. Unfortunately, mahanimbine is available with few commercial sources at very high prices. Therefore, we prepared a curry leaves extract and isolated mahanimbine by a novel, yet simple, extraction method that requires less time and causes minimum environmental hazards. After purification, structure, and mass were confirmed for the isolated compound by IR spectrum and LC-MS/MS analysis, respectively. In the mechanistic study, hydrolysis of ATP and substrate efflux by P-gp are coupled. Hence, ATP binding at the ATPase-binding site is one of the fundamental steps for the P-gp efflux cycle. We found that mahanimbine demonstrated to stimulate P-gp ATPase activity. Concurrently, it enhanced the intracellular accumulation of P-gp substrates Rhodamine 123 and Hoechst stain, which indicates that mahanimbine modulates the function of P-gp. In addition, we have analyzed the complementary effect of mahanimbine with the chemotherapeutic drug gefitinib. We found that mahanimbine synergistically enhanced gefitinib efficiency by increasing its intracellular accumulation in lung cancer cells. Overall, mahanimbine has been shown to be a potent P-gp modulator. Therefore, mahanimbine can be further developed as a potential candidate to overcome chemoresistance in lung cancer.

Application of a high-resolution in vitro human MDR1-MDCK assay and in vivo studies in preclinical species to improve prediction of CNS drug penetration.

P-glycoprotein (P-gp, MDR1) is expressed at the blood-brain barrier (BBB) and restricts penetration of its substrates into the central nervous system (CNS). In vitro MDR1 assays are frequently used to predict the in vivo relevance of MDR1-mediated efflux at the BBB. It has been well established that drug candidates with high MDR1 efflux ratios (ERs) display poor CNS penetration. Following a comparison of MDR1 transporter function between the MDR1-MDCKI cell line from National Institutes of Health (NIH) and our internal MDR1-MDCKII cell line, the former was found to provide better predictions of in vivo brain penetration than our in-house MDR1-MDCKII cell line. In particular, the NIH MDR1 assay has an improved sensitivity to differentiate the compounds with ERs of <3 in our internal cell line and is able to reduce the risk of false negatives. A better correlation between NIH MDR1 ERs and brain penetration in rat and non-human primate (NHP) was demonstrated. Additionally, a comparison of brain penetration time course of MDR1 substrates and an MDR1 non-substrate in NHP demonstrated that MDR1 interaction can delay the time to equilibrium of drug concentration in the brain with plasma. It is recommended to select highly permeable compounds without MDR1 interaction for rapid brain penetration to produce the maximal pharmacological effect in the CNS with a quicker onset.

Screening novel CNS drug candidates for P-glycoprotein interactions using the cell line iP-gp: In vitro efflux ratios from iP-gp and MDCK-MDR1 monolayers compared to brain distribution data from mice.

Drug efflux by P-glycoprotein (P-gp, ABCB1) is considered as a major obstacle for brain drug delivery for small molecules. P-gp-expressing cell monolayers are used for screening of new drug candidates during early states of drug development. It is, however, uncertain how well the in vitro studies can predict the in vivo P-gp mediated efflux at the blood-brain barrier (BBB). We previously developed a novel cell line of porcine origin, the iP-gp cell line, with high transepithelial resistance and functional expression of human P-gp. The aim of the present study was to evaluate the applicability of the cell line for screening of P-gp interactions of novel drug candidates. For this purpose, bidirectional fluxes of 14 drug candidates were measured in iP-gp cells and in MDCK-MDR1 cells, and compared with pharmacokinetic data obtained in male C57BL/6 mice. The iP-gp cells formed extremely tight monolayers (>15 000 Ω∙cm2) as compared to the MDCK- MDR1 cells (>250 Ω∙cm2) and displayed lower Papp,a-b values. The efflux ratios obtained with iP-gp and MDCK-MDR1 monolayers correlated with Kp,uu,brain values from the in vivo studies, where compounds with the lowest Kp,uu,brain generally displayed the highest efflux ratios. 12 of the tested compounds displayed a poor BBB penetration in mice as judged by Kp,uu less than 1. Of these compounds, nine compounds were categorized as P-gp substrates in the iP-gp screening, whereas analysis of data estimated in MDCK-MDR1 cells indicated four compounds as potential substrates. The results suggest that the iP-gp cell model may be a sensitive and useful screening tool for drug screening purposes to identify possible substrates of human P-glycoprotein.

Reveals of candidate active ingredients in Justicia and its anti-thrombotic action of mechanism based on network pharmacology approach and experimental validation.

Thrombotic diseases seriously threaten human life. Justicia, as a common Chinese medicine, is usually used for anti-inflammatory treatment, and further studies have found that it has an inhibitory effect on platelet aggregation. Therefore, it can be inferred that Justicia can be used as a therapeutic drug for thrombosis. This work aims to reveal the pharmacological mechanism of the anti-thrombotic effect of Justicia through network pharmacology combined with wet experimental verification. During the analysis, 461 compound targets were predicted from various databases and 881 thrombus-related targets were collected. Then, herb-compound-target network and protein-protein interaction network of disease and prediction targets were constructed and cluster analysis was applied to further explore the connection between the targets. In addition, Gene Ontology (GO) and pathway (KEGG) enrichment were used to further determine the association between target proteins and diseases. Finally, the expression of hub target proteins of the core component and the anti-thrombotic effect of Justicia's core compounds were verified by experiments. In conclusion, the core bioactive components, especially justicidin D, can reduce thrombosis by regulating F2, MMP9, CXCL12, MET, RAC1, PDE5A, and ABCB1. The combination of network pharmacology and the experimental research strategies proposed in this paper provides a comprehensive method for systematically exploring the therapeutic mechanism of multi-component medicine.

Antioxidant effect of ascorbic acid against cisplatin-induced nephrotoxicity and P-glycoprotein expression in rats.

Cisplatin (CDDP) is a highly potent anticancer drug that is widely used in the treatment of several cancers. CDDP-induced nephrotoxicity (CIN) is one of the most significant adverse effects, and oxidative stress is thought to be one of the mechanisms underlying CIN. Although there are some studies available on the variability in transporter expression in the kidney after a single CDDP dose, none have reported the change in renal transporter expression after multiple CDDP dose administrations. P-glycoprotein (P-gp), a transporter, is reported to be induced by oxidative stress. Ascorbic acid is a vitamin with antioxidant potential and therefore, may regulate the expression of P-gp transporter and affect CIN. In the present study, our aim was to assess the variability in expression of several renal transporters after multiple CDDP dose administrations and the antioxidant effect of ascorbic acid against transporter expression and CIN. Multiple doses of CDDP affected markers of kidney injury and antioxidants in the kidneys. Also, the expression of P-gp, breast cancer resistance protein, and multidrug resistance-associated protein 4 was upregulated by CDDP. Using a normal kidney cell line, we demonstrated that ascorbic acid attenuated CDDP-induced cytotoxicity due to its high superoxide scavenging ability. CDDP and ascorbic acid were injected into rats once a week for three weeks, and it was observed that co-administration of ascorbic acid attenuated CIN and regulated antioxidant marker. In addition, ascorbic acid reduced P-gp expression, which was upregulated by CDDP. In conclusion, ascorbic acid may attenuate CIN and reverse P-gp-mediated changes in drug pharmacokinetics.

Isobavachalcone as an Active Membrane Perturbing Agent and Inhibitor of ABCB1 Multidrug Transporter.

Isobavachalcone (IBC) is an active substance from the medicinal plant Psoralea corylifolia. This prenylated chalcone was reported to possess antioxidative, anti-inflammatory, antibacterial, and anticancer activities. Multidrug resistance (MDR) associated with the over-expression of the transporters of vast substrate specificity such as ABCB1 (P-glycoprotein) belongs to the main causes of cancer chemotherapy failure. The cytotoxic, MDR reversing, and ABCB1-inhibiting potency of isobavachalcone was studied in two cellular models: human colorectal adenocarcinoma HT29 cell line and its resistant counterpart HT29/Dx in which doxorubicin resistance was induced by prolonged drug treatment, and the variant of MDCK cells transfected with the human gene encoding ABCB1. Because MDR modulators are frequently membrane-active substances, the interaction of isobavachalcone with model phosphatidylcholine bilayers was studied by means of differential scanning calorimetry. Molecular modeling was employed to characterize the process of membrane permeation by isobavachalcone. IBC interacted with ABCB1 transporter, being a substrate and/or competitive inhibitor of ABCB1. Moreover, IBC intercalated into model membranes, significantly affecting the parameters of their main phospholipid phase transition. It was concluded that isobavachalcone interfered both with the lipid phase of cellular membrane and with ABCB1 transporter, and for this reason, its activity in MDR cancer cells was presumptively beneficial.

Quantification of P-Glycoprotein in the Gastrointestinal Tract of Humans and Rodents: Methodology, Gut Region, Sex, and Species Matter.

Intestinal efflux transporters affect the gastrointestinal processing of many drugs but further data on their intestinal expression levels are required. Relative mRNA expression and relative and absolute protein expression data of transporters are commonly measured by real-time polymerase chain reaction (RT-PCR), Western blot and mass spectrometry-based targeted proteomics techniques. All of these methods, however, have their own strengths and limitations, and therefore, validation for optimized quantification methods is needed. As such, the identification of the most appropriate technique is necessary to effectively translate preclinical findings to first-in-human trials. In this study, the mRNA expression and protein levels of the efflux transporter P-glycoprotein (P-gp) in jejunal and ileal epithelia of 30 male and female human subjects, and the duodenal, jejunal, ileal and colonic tissues in 48 Wistar rats were quantified using RT-PCR, Western blot and liquid chromatography-tandem mass spectrometry (LC-MS/MS). A similar sex difference was observed in the expression of small intestinal P-gp in humans and Wistar rats where P-gp was higher in males than females with an increasing trend from the proximal to the distal parts in both species. A strong positive linear correlation was determined between the Western blot data and LC-MS/MS data in the small intestine of humans (R2 = 0.85). Conflicting results, however, were shown in rat small intestinal and colonic P-gp expression between the techniques (R2 = 0.29 and 0.05, respectively). In RT-PCR and Western blot, an internal reference protein is experimentally required; here, beta-actin was used which is innately variable along the intestinal tract. Quantification via LC-MS/MS can provide data on P-gp expression without the need for an internal reference protein and consequently, can give higher confidence on the expression levels of P-gp along the intestinal tract. Overall, these findings highlight similar trends between the species and suggest that the Wistar rat is an appropriate preclinical animal model to predict the oral drug absorption of P-gp substrates in the human small intestine.

Antibacterial Activity and Reversal of Multidrug Resistance of Tumor Cells by Essential Oils from Fresh Leaves, Flowers, and Stems of Montanoa quadrangularis Schultz Bipontinus (Asteraceae) Collected in Mérida-Venezuela.

Essential oils obtained by hydrodistillation of Montanoa quadrangularis leaves, flowers, and stems were analyzed by GC and GC/MS techniques revealing myrcene, limonene, ß-phellandrene, and sabinene among the main components. The aim of the present study was to evaluate the MDR modulator activity on human MDR1 gene transfected mouse lymphoma cell line and the antimicrobial activity on the essential oils obtained from different parts of the species under investigation. The results revealed that MQL caused a similar increase in the fluorescence activity of the cells at 0.02 µL/mL comparing to the Verapamil® value. The antimicrobial assay was carried out according to the disc diffusion method. Five different bacterial strains (Staphylococcus epidermidis, Bacillus subtilis, Pseudomonas aeruginosa, Escherichia coli AG 100, and Escherichia coli AG100A) were treated with the essential oils and the zones of inhibition were determined on TSA plates and TSA agar plates supplemented with Tween 20. MQF and MQL showed activity against B. subtilis, S. epidermidis, and E. coli AG 100A while MQS was only active against E. coli AG 100A on TSA agar plates experiment. In case of TSA agar plates supplemented with 0.1 v/v% Tween 20 detergent, MQF showed inhibition on B. subtilis, S. epidermidis, and E. coli AG 100A; MQL was active against B. subtilis, E. coli AG 100, and E. coli AG 100A while MQS was only active against E. coli AG 100A.

Interaction of Phytocompounds of Echinacea purpurea with ABCB1 and ABCG2 Efflux Transporters.

Preparations of Echinacea purpurea (E. purpurea) are widely used for the management of upper respiratory infections, influenza, and common cold, often in combination with other conventional drugs. However, the potential of phytochemical constituents of E. purpurea to cause herb-drug interactions via ABCB1 and ABCG2 efflux transporters remains elusive. The purpose of this study was to investigate the impact of E. purpurea-derived caffeic acid derivatives (cichoric acid and echinacoside) and tetraenes on the mRNA and protein expression levels as well as on transport activity of ABCB1 and ABCG2 in intestinal (Caco-2) and liver (HepG2) cell line models. The safety of these compounds was investigated by estimating EC20 values of cell viability assays in both cell lines. Regulation of ABCB1 and ABCG2 protein in these cell lines were analyzed after 24 h exposure to the compounds at 1, 10, and 50 µg/mL. Bidirectional transport of 0.5 µg/mL Hoechst 33342 and 5 µM rhodamine across Caco-2 monolayer and profiling for intracellular concentrations of the fluorophores in both cell lines were conducted to ascertain inhibition effects of the compounds. Cichoric acid showed no cytotoxic effect, while the EC20 values of tetraenes and echinacoside were 45.0 ± 3.0 and 52.0 ± 4.0 µg/mL in Caco-2 cells and 28.0 ± 4.3 and 62.0 ± 9.9 µg/mL in HepG2 cells, respectively. In general, the compounds showed heterogeneous induction of ABCB1 with the strongest 3.6 ± 1.2-fold increase observed for 10 µg/mL tetraenes in Caco-2 cells (p < 0.001). However, the compounds did not induce ABCG2. None of the phytocompounds inhibited significantly net flux of the fluorophores across Caco-2 monolayers. Overall, tetraenes moderately induced ABCB1 but not ABCG2 in Caco-2 and HepG2 cells while no compound significantly inhibited activity of these transporters at clinically relevant concentration to cause herb-drug interactions.

Poncirin downregulates ATP-binding cassette transporters to enhance cisplatin sensitivity in cisplatin-resistant osteosarcoma cells.

Poncirin, a flavanone glycoside with bitter taste extracted from dried immature fruit of Poncirus trifoliate, exhibits multiple biological activities including anti-tumor activity. Our study aimed to determine the effect and potential mechanism of poncirin on cisplatin resistance in osteosarcoma (OS) cells. CCK-8, flow cytometry analysis, and caspase-3/7 activity assays were used to evaluate cisplatin sensitivity. The expression changes of multidrug resistance 1 (MDR1), multidrug resistance-associated protein (MRP1), breast cancer resistance protein (BCRP), and phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) pathway-related proteins were detected by RT-qPCR or western blot analyses. Results showed that poncirin exposure enhanced cisplatin sensitivity, promoted apoptosis, and increased caspase-3/7 activity in cisplatin-resistant OS cells. Poncirin decreased the expression levels of MDR1, MRP1, and BCRP, and inhibited the PI3K/Akt signaling in OS cells. Rescue experiments suggested that activation of the PI3K/Akt signaling by 740Y-P abolished poncirin-induced expression reduction of MDR1, MRP1, and BCRP, and attenuated the facilitative effects of poncirin on cisplatin sensitivity and apoptosis in cisplatin-resistant OS cells. In summary, poncirin suppressed cisplatin resistance in cisplatin-resistant OS cells by downregulating the expression of MDR1, MRP1, and BCRP through inhibiting the PI3K/Akt pathway.

Overcoming vincristine resistance in cancer: Computational design and discovery of piperine-inspired P-glycoprotein inhibitors.

P-glycoprotein (P-gp)/MDR-1 plays a major role in the development of multidrug resistance (MDR) by pumping the chemotherapeutic drugs out of the cancer cells and reducing their efficacy. A number of P-gp inhibitors were reported to reverse the MDR when co-administered with chemotherapeutic drugs. Unfortunately, none has approved for clinical use due to toxicity issues. Some of the P-gp inhibitors tested in the clinics are reported to have cross-reactivity with CYP450 drug-metabolizing enzymes, resulting in unpredictable pharmacokinetics and toxicity of co-administered chemotherapeutic drugs. In this study, two piperine analogs (3 and 4) having lower cross-reactivity with CYP3A4 drug-metabolizing enzyme are identified as P-glycoprotein (P-gp) inhibitors through computational design, followed by synthesis and testing in MDR cancer cell lines over-expressing P-gp (KB ChR 8-5, SW480-VCR, and HCT-15). Both the analogs significantly increased the vincristine efficacy in MDR cancer cell lines at low micromole concentrations. Specifically, 3 caused complete reversal of vincristine resistance in KB ChR 8-5 cells and found to act as competitive inhibitor of P-gp as well as potentiated the vincristine-induced NF-KB-mediated apoptosis. Therefore, 3 ((2E,4E)-1-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)-5-(4-hydroxy-3-methoxyphenyl)penta-2,4-dien-1-one) can serve as a potential P-gp inhibitor for in vivo investigations, to reverse multidrug resistance in cancer.

Plant extracts and betulin from Ligaria cuneifolia inhibit P-glycoprotein function in leukemia cells.

Overexpression of P-glycoprotein (P-gp), which is linked to multidrug resistance (MDR), is one of the underlying obstacles to the success of chemotherapy as it reduces the efficacy of anticancer drugs and the side effects of these increase as a result of any increased dose to achieve the therapeutic effect. To identify agents with P-gp inhibitory properties, ethanol extracts from 80 plants were screened for their ability to increase intracellular doxorubicin-associated fluorescence, and the extract of Ligaria cuneifolia was found to be the most effective. Its bioassay-guided isolation yielded the pentacyclic triterpene betulin as active agent. This efficiently inhibited P-gp mediated efflux, as demonstrated by the enhancement of the intracellular accumulation of doxorubicin and rhodamine 123 from 1.56 µM in the P-gp overexpressing MDR leukemia cell, Lucena 1. Betulin was also able to render Lucena 1 sensitive to Dox from 0.39 µM. The docking studies revealed that betulin tightly binds to a key region of the TMDs, with a binding mode overlapping one main site of doxorubicin and, more interestingly, emulating the same contacts as tariquidar, as revealed by the per-residue energetic analysis from molecular dynamics simulations. MTT assay using peripheral blood mononuclear cells and hemolysis assay showed that betulin is devoid of toxicity. These findings provide important evidence that betulin may be a safe and promising entity to be further investigated to develop agents able to overcome P-gp-mediated MDR, resulting in a more effective and less toxic chemotherapy.

Da-Huang-Xiao-Shi decoction protects against3, 5-diethoxycarbonyl-1,4-dihydroxychollidine-induced chronic cholestasis by upregulating bile acid metabolic enzymes and efflux transporters.

ETHNOPHARMACOLOGICAL RELEVANCE: Chronic cholestasis is a usual clinical pathological process in hepatopathy and has few treatment options; it is classified under the category of jaundice in Chinese medicine. Da-Huang-Xiao-Shi decoction (DHXSD) is a classic Chinese prescription which is used to treat jaundice. AIM OF THE STUDY: We aimed to examine the protective effect of DHXSD on liver and its potential mechanism of action against chronic cholestasis. MATERIALS AND METHODS: Chronic cholestasis was induced using 3, 5-diethoxycarbonyl-1,4-dihydroxychollidine (DDC) in mice. Mice were then administered DHXSD intragastrically at doses of 3.68, 7.35, and 14.70 g/kg for four weeks followed by further analyses. Serum biochemical indices and liver pathology were explored. Eighteen individual bile acids (BAs) in mice serum and liver were quantified using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The expression of BA related metabolic enzymes, transporters, along with nuclear receptor farnesoid X receptor (FXR) was detected by real-time qPCR and Western blot. RESULTS: DHXSD treatment reduced the serum biochemical indices, ameliorated pathological injury, and improved the disordered BA homeostasis. Mice treated with DHXSD showed significantly upregulated expression of the metabolic enzymes, cytochrome P450 2b10 (Cyp2b10), Cyp3a11, and UDP-glucuronosyltransferase 1a1 (Ugt1a1); and the bile acid transporters, multidrug resistance protein 2 (Mdr2), bile salt export pump (Bsep), and multidrug resistance-associated protein 3 (Mrp3). DHXSD treatment also significantly upregulated FXR expression in mice with DDC-induced chronic cholestasis. CONCLUSIONS: DHXSD exerted protective effects on chronic cholestasis in DDC-treated mice by alleviating the disordered homeostasis of BAs through increased expression of BA related metabolic enzymes and efflux transporters.

The Intestinal Efflux Transporter Inhibition Activity of Xanthones from Mangosteen Pericarp: An In Silico, In Vitro and Ex Vivo Approach.

The capacity of α-mangostin (α-MG) and ß-mangostin (ß-MG) from mangosteen pericarp on P-glycoprotein (Pgp) in silico, in vitro, and ex vivo was investigated in this study. Screening with the ADMET Predictor™ program predicted the two compounds to be both a Pgp inhibitor and Pgp substrate. The compounds tended to interact with Pgp and inhibit Pgp ATPase activity. Additionally, bidirectional transport on Caco-2 cell monolayers demonstrated a significantly lower efflux ratio than that of the control (α-(44.68) and ß-(46.08) MG versus the control (66.26); p < 0.05) indicating an inhibitory effect on Pgp activity. Test compounds additionally revealed a downregulation of MDR1 mRNA expression. Moreover, an ex vivo absorptive transport in everted mouse ileum confirmed the previous results that α-MG had a Pgp affinity inhibitor, leading to an increase in absorption of the Pgp substrate in the serosal side. In conclusion, α- and ß-MG have the capability to inhibit Pgp and they also alter Pgp expression, which makes them possible candidates for reducing multidrug resistance. Additionally, they influence the bioavailability and transport of Pgp substrate drugs.