Down-regulation Of P-glycoprotein Research Articles

Overcoming Breast Cancer Drug Resistance: A Novel Approach Using siRNA-Mediated P-glycoprotein Downregulation to Enhance Vinorelbine Efficacy.

Cancer, the second leading cause of mortality worldwide, represents a global health challenge, primarily due to drug resistance. Vinorelbine is a chemotherapeutic agent that disrupts cancer cell growth by targeting microtubules and inducing apoptosis. However, drug resistance remains a formidable obstacle. This resistance is caused by various factors including genetic mutations, drug efflux mechanisms, and DNA repair systems. Resolution of this challenge requires an innovative approach. This study investigated the potential of small interfering RNA (siRNA) to target and downregulate a vinorelbine-resistant MCF-7/ADR breast cancer cell line. Cells were cultured in Dulbecco's modified Eagle's medium (DMEM) 10% fetal bovine serum/penicillin/streptomycin. An siRNA targeting ABCB1 was designed and synthesized, and the cells were transfected with siRNA at final concentrations of 10, 20, and 30 nM. The3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to assess cell viability. ABCB1 mRNA expression levels were determined by real-time polymerase chain reaction (PCR). MCF-7 cells exhibited a higher sensitivity to vinorelbine than MCF-7/ADR cells. MCF-7/ADR cells exhibited resistance to vinorelbine at concentrations, 12.50 and 25.00 μM. Treatment with siRNA significantly reduced ABCB1 expression by 2.93-fold (P=0.0001). Similarly, co-treatment with siRNA and vinorelbine produced a substantial 2.89-fold decrease in ABCB1 gene expression in MCF-7 cells compared to that in MCF-7/ADR cells (P=0.0001). The results of the present study indicate that the concurrent use of siRNA and vinorelbine holds substantial promise as a therapeutic approach to overcome ABCB1-mediated multidrug resistance (MDR) in breast cancer. It is necessary to conduct comprehensive clinical trials to determine the true effectiveness of this combination therapy.

Overcoming Multidrug Resistance by Base-Editing-Induced Codon Mutation.

Multidrug resistance (MDR) is the main obstacle in cancer chemotherapy. ATP binding cassette (ABC) transporters on the MDR cell membrane can transport a wide range of antitumor drugs out of cells, which is one of the main causes of MDR. Therefore, disturbing ABC transporters becomes the key to reversing MDR. In this study, we implement a cytosine base editor (CBE) system to knock out the gene encoding ABC transporters by base editing. When the CBE system works in MDR cells, the MDR cells are manipulated, and the genes encoding ABC transporters can be inactivated by precisely changing single in-frame nucleotides to induce stop (iSTOP) codons. In this way, the expression of ABC efflux transporters is reduced and intracellular drug retention is significantly increased in MDR cells. Ultimately, the drug shows considerable cytotoxicity to the MDR cancer cells. Moreover, the substantial downregulation of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) implies the successful application of the CBE system in the knockout of different ABC efflux transporters. The recovery of chemosensitivity of MDR cancer cells to the chemotherapeutic drugs revealed that the system has a satisfactory universality and applicability. We believe that the CBE system will provide valuable clues for the use of CRISPR technology to defeat the MDR of cancer cells.

Cutting Off H+ Leaks on the Inner Mitochondrial Membrane: A Proton Modulation Approach to Selectively Eradicate Cancer Stem Cells.

Cancer stem cells (CSCs) are associated with the invasion and metastatic relapse of various cancers. However, current cancer therapies are limited to targeting the bulk of primary tumor cells while remaining the CSCs untouched. Here, we report a new proton (H+) modulation approach to selectively eradicate CSCs via cutting off the H+ leaks on the inner mitochondrial membrane (IMM). Based on the fruit extract of Gardenia jasminoides, a multimodal molecule channel blocker with high biosafety, namely, Bo-Mt-Ge, is developed. Importantly, in this study, we successfully identify that mitochondrial uncoupling protein UCP2 is closely correlated with the stemness of CSCs, which may offer a new perspective for selective CSC drug discovery. Mechanistic studies show that Bo-Mt-Ge can specifically inhibit the UCP2 activities, decrease the H+ influx in the matrix, regulate the electrochemical gradient, and deplete the endogenous GSH, which synergistically constitute a unique MoA to active apoptotic CSC death. Intriguingly, Bo-Mt-Ge also counteracts the therapeutic resistance via a two-pronged tactic: drug efflux pump P-glycoprotein downregulation and antiapoptotic factor (e.g., Bcl-2) inhibition. With these merits, Bo-Mt-Ge proved to be one of the safest and most efficacious anti-CSC agents, with ca. 100-fold more potent than genipin alone in vitro and in vivo. This study offers new insights and promising solutions for future CSC therapies in the clinic.

Reduction of breast tumor drug resistance by 2,3,5,4'-tetrahydroxystilbene for exhibition synergic chemotherapeutic effect.

Chemotherapy drugs have limited efficacy in breast cancer due to multidrug resistance generated by cancer cells against anticancer drugs. In this study, we developed a novel derivative, 2, 3, 5, 4‘-tetrahydroxystilbene (TG1) by modifying 2, 3, 5, 4‘-tetrahydroxystilbene-2-O-beta-D-glucoside (THSG). In-vivo zebrafish embryo tests revealed that TG1 showed low toxicity. The equitoxic combination of DOX or DTX with TG1 in MCF-7/Adr reduced the IC50 of DOX or DTX, and the combination index (CI) showed strong synergistic effects in the 1:3 molar ratio of DTX: TG1 and 1:5 molar ratio of DOX: TG1. Moreover, fluorescence images confirmed the cellular uptake of DOX when combined with TG1 in MCF-7/Adr. Western blotting analysis indicated downregulation of p-glycoprotein (P-gp) after MCF-7/Adr treated with TG1. In conclusion, the combined therapy of DTX or DOX and TG1 increases drug efficacy via suppressing the p-glycoprotein efflux pump. These results suggest that TG1 may have potential use for breast cancer patients, especially those with multidrug resistance.

A novel targeted co-delivery nanosystem for enhanced ovarian cancer treatment via multidrug resistance reversion and mTOR-mediated signaling pathway

BackgroundMultidrug resistance (MDR) is the main challenge of successful chemotherapy for ovarian cancer patients, with 50% to 75% of ovarian cancer patients eventually relapsed due to it. One of the effective strategies for treating MDR and improving therapeutic efficiency of ovarian cancer is to use nanotechnology-based targeted drug delivery systems. In this study, a novel nano targeted co-delivery system modified by hyaluronic acid (HA) was developed by using gold nanorods coated with functionalized mesoporous silica nanoparticles (HA-PTX/let-7a-GNR@MSN) for combined delivery of hydrophobic chemotherapy drug Paclitaxel (PTX) and lethal-7a (let-7a), a microRNA (miR), to overcome MDR in ovarian cancer. Furthermore, we also analyzed the molecular mechanism of this nanotherapeutic system in the treatment of ovarian cancer.ResultsHA-modified nanocomplexes can specifically bind to the CD44 receptor, which is highly expressed in SKOV3/SKOV3TR cells, achieving effective cell uptake and 150% enhancement of tumor site permeability. The nanosystem realized the stable combination and protective transportation of PTX and miRs. Analysis of drug-resistant SKOV3TR cells and an SKOV3TR xenograft model in BALB/c-nude mice showed significant downregulation of P-glycoprotein in heterogeneous tumor sites, PTX release, and subsequent induction of apoptosis. More importantly, this nanosystem could synergistically inhibit the growth of ovarian tumors. Further studies suggest that mTOR-mediated signaling pathways play an important role in reversing drug resistance and inducing apoptosis.ConclusionsTo sum up, these data provide a model for overcoming PTX resistance in ovarian cancer.Graphical

Rhinacanthin-C enhances chemosensitivity of breast cancer cells via the downregulation of P-glycoprotein through inhibition of Akt/NF-kappa B signaling pathway

Introduction: High expression of P-glycoprotein (P-gp) has been linked to multidrug resistance (MDR) and chemotherapeutic failure. Previously, we demonstrated that rhinacanthin-C, a naphthoquinone from Rhinacanthus nasutus, was able to enhance the cytotoxicity of doxorubicin against breast cancer MCF-7 cells via direct P-gp inhibition. In this study, we looked at its effect on P-gp downregulation and the mechanism involved in the resistance of MCF-7 cells to doxorubicin. Methods: Doxorubicin-resistant MCF-7 (MCF-7/DOX) cells were exposed to rhinacanthin-C for 24-48 hours prior to the assessment of their chemosensitivity via MTT assay, P-gp activity via calcein-AM uptake assay, P-gp expression, and signaling via qRT-PCR and western blot analyses. Results: Pretreatment with 1 µM of rhinacanthin-C for 48 hours significantly enhanced cytotoxicity of doxorubicin, as well as camptothecin and etoposide, to MCF-7/DOX cells. In the rhinacanthin-C-treated cells, reduction of MDR1 mRNA and P-gp levels and increased intracellular calcein were observed. Moreover, phosphorylation of Akt, NF-ᴋB and IκB-α, along with YB-1 expression, significantly decreased after 24-hour treatment with rhinacanthin-C. In contrast, the naphthoquinone had no effect on expression levels of ERK1/2 and phosphorylated ERK1/2 under similar conditions. Conclusion: Rhinacanthin-C, at a non-cytotoxic concentration (1 µM), could downregulate P-gp expression in MCF-7/DOX cells via the inhibition of the Akt/NF-ᴋB signaling pathway and YB-1 expression. Long-term exposure to this natural naphthoquinone may increase the chemosensitivity of cancer cells with MDR phenotype.

The study of Raddeanin A cerebrovascular endothelial cell trafficking through P-glycoprotein

As one of the natural triterpenoids isolated from Anemone Raddeana Regel, Raddeanin A (RA) has been confirmed to possess therapeutic effects against multiple tumorigeneses, especially for the onset of glioblastoma and growth in human brains. However, the mechanism by which this happens remains poorly understood in terms of the vascular endothelium trafficking routine of RA through the brain-blood barrier (BBB). To seek such answers, human brain microenvironment endothelial cells (HBMECs) were used to stimulate the microenvironment in vitro, and to explore the intracellular accumulation of RA. The results of this experiment illustrated that RA has a relative moderate transport affinity for such cells. The kinetic parameter Km was 37.01 ± 2.116 μM and Vmax was 9.412 ± 0.1375 nM/min/mg of protein. Interestingly, protein downregulation of P-glycoprotein (P-gp, ABCB1/MDR1) significantly activated RA transmembrane activity, which proves that P-gp is responsible for RA cellular trafficking. In addition, the selective non-specific inhibitor, LY335979 increased either RA or the classical substrate of P-gp, digoxin, intracellular accumulation by restricting the transporter's function but without jeopardizing cytomembrane proteins. Moreover, a decrease in the expression or activity of P-gp triggered RA drug resistance to HBMECs. In summary, our data showed that both the expression and function of P-gp are all necessary for RA transmembrane trafficking through cerebrovascular endothelial cells. This study provides significant evidence for the presence of a connection between RA transport and P-gp variation during drug BBB penetration. It is also suggesting some vital guidance on the RA pharmacodynamic effect in human brains.

A Novel Targeted Co-Delivery Nanosystem for Enhanced Ovarian Cancer Treatment Via Multidrug Resistance Reversion

Multidrug resistance (MDR) is a major challenge in successful chemotherapy treatment of ovarian cancer patients, and 50%–75% of ovarian cancer patients eventually relapse because of MDR. One of an effective strategy for treating MDR and improving therapeutic efficiency of ovarian cancer is to use nanotechnology-based targeted drug delivery systems. This study developed a novel hyaluronic acid (HA)-targeted co-delivery nanosystem using functionalized mesoporous silica nanoparticle–coated gold nanorods (HA-PTX/let-7a-GNR@MSN) to co-deliver paclitaxel (PTX), a hydrophobic chemotherapy drug, and lethal-7a (let-7a), a microRNA (miR), to overcome MDR in ovarian cancer. We also analyzed the molecular mechanism of miR let-7a in ovarian cancer treatment. The nanosystem enabled protective drug delivery and stable binding of PTX and miRs. Analysis of drug-resistant SKOV3TR cells and an SKOV3TR xenograft model in BALB/c-nude mice showed significant P-glycoprotein downregulation in heterogeneous tumor sites, PTX release, and apoptosis induction later. Results showed that HA-modified nanocomposites can specifically bind to the CD44 receptor, which is highly expressed in SKOV3/SKOV3 TR cells, achieving effective cell uptake and 150% enhancement of tumor site permeability. More importantly, this nanosystem resulted in synergistic inhibition of ovarian tumor growth. Overall, the data provide a model for overcoming PTX resistance in ovarian cancer.

Emodin alleviates gemcitabine resistance in pancreatic cancer by inhibiting MDR1/P-glycoprotein and MRPs expression.

Gemcitabine is a gold standard chemotherapeutic agent for pancreatic cancer. However, gemcitabine has limited effectiveness due to the short-term development of chemoresistance. Emodin, a natural anthraquinone derivative isolated from the roots of rheumatic palm leaves prevents immunosuppression and exerts anticancer effects. The present study aimed to evaluate the effect of emodin on gemcitabine resistance. Gemcitabine-resistant PANC-1 pancreatic cancer cell xenografts were established in athymic mice, which were randomly assigned into four treatments groups as follows: Gemcitabine group, Emodin group, Gemcitabine+Emodin group and Negative control group. Body weight, tumor volume and tumor weight were measured over the course of treatment. The effect of each treatment on tumor tissue proliferation and apoptosis from nude mice was evaluated by using immunohistochemistry. The effect of each treatment on the proliferation of gemcitabine-resistant PANC-1 cells was also determined by using the Cell Counting Kit-8. Then, reverse transcription-quantitative (RT-q) PCR and western blotting were used to detect the mRNA and protein expression, respectively, of multidrug resistance gene 1 (MDR1) and the drug resistance-related proteins MRP1 and MRP5. The function and expression level of DR1 gene product, p-glycoprotein, was also analysed by flow cytometry and RT-qPCR, respectively. The results demonstrated that the combination of gemcitabine and emodin significantly reduced xenograft volume and reduced tumor growth in mice compared with treatment with gemcitabine or emodin only. In addition, emodin treatment reduced resistance to gemcitabine, which was characterized by the downregulation of P-glycoprotein, MRP1 and MRP5 expression in the group receiving combination treatment. The level of P-glycoprotein was also decreased in the group treated with gemcitabine+emodin compared with the single treatment groups. Taken together, these results demonstrated that emodin enhanced gemcitabine efficacy in tumor treatment and alleviated gemcitabine resistance in PANC-1 cell xenografts in mice via suppressing MDR1/P-glycoprotein and MRP expression.

A Co-Delivery System of Curcumin and p53 for Enhancing the Sensitivity of Drug-Resistant Ovarian Cancer Cells to Cisplatin.

In order to enhance the sensitivity of drug-resistant ovarian cancer cells to cisplatin (DDP), a co-delivery system was designed for simultaneous delivery of curcumin (CUR) and p53 DNA. Firstly, the bifunctional peptide K14 composed of tumor targeting peptide (tLyP-1) and nuclear localization signal (NLS) was synthesized. A nonviral carrier (PEI-K14) was synthesized by cross-linking low molecular weight polyethyleneimine (PEI) with K14. Then, CUR was coupled to PEI-K14 by matrix metalloproteinase 9 (MMP9)-cleavable peptide to prepare CUR-PEI-K14. A co-delivery system, named CUR-PEI-K14/p53, was obtained by CUR-PEI-K14 and p53 self-assembly. Furthermore, the physicochemical properties and gene transfection efficiency were evaluated. Finally, ovarian cancer cisplatin-resistant (SKOV3-DDP) cells were selected to evaluate the effect of CUR-PEI-K14/p53 on enhancing the sensitivity of drug-resistant cells to DDP. The CUR-PEI-K14/DNA complexes appeared uniformly dispersed and spherical. The particle size was around 20–150 nm and the zeta potential was around 18–37 mV. It had good stability, high transfection efficiency, and low cytotoxicity. CUR-PEI-K14/p53 could significantly increase the sensitivity of SKOV3-DDP cells to DDP, and this effect was better as combined with DDP. The sensitizing effect might be related to the upregulation of p53 messenger RNA (mRNA), the downregulation of P-glycoprotein (P-gp) mRNA, and the upregulation of BCL2-Associated X (bax) mRNA. CUR-PEI-K14/p53 can be used as an effective strategy to enhance the sensitivity of drug-resistant ovarian cancer cells to DDP.

Diminished Expression of P-glycoprotein Using Focused Ultrasound Is Associated With JNK-Dependent Signaling Pathway in Cerebral Blood Vessels

MRI-guided focused ultrasound (MRgFUS) combined with microbubbles (MBs) is a promising technology that can facilitate drug delivery through a temporarily disrupted blood-brain barrier (BBB) and induce the down-regulation of P-glycoprotein (P-gp) expression on the blood vessels. Despite the increasing evidence regarding the down-regulation of P-gp expression after MRgFUS BBB disruption (BBBD), its underlying molecular events remain unclear. The aim of this study was to evaluate the underlying mechanism of FUS BBBD-mediated P-gp down-regulation. While our results showed down-regulation of P-gp at 24 h post-BBBD in transcriptional and translational levels, restoration to the normal expression appeared at different time points for transcriptional (72 h) and translational (120 h) levels. In addition, the signaling molecule, JNK, was significantly activated in the cerebral blood vessels at 24 h post-BBBD. Although P-gp levels were significantly decreased, the expression levels of proteins involved in the integrity of blood vessels, such as Glut1, ZO-1 and occludin, were not decreased at 24 h post-BBBD. Our study suggests that the JNK signaling pathway is involved in the regulation of FUS-induced P-gp expression, without affecting vessel integrity, and a detailed regulatory mechanism can provide the basis for clinical application of FUS to the treatment of neurological disease.

Administration of Vitamin D Metabolites Affects RNA Expression of Xenobiotic Metabolising Enzymes and Function of ABC Transporters in Rats

From studies on different species and in cell culture systems, it has been suggested that vitamin D metabolites might affect the metabolism and elimination of xenobiotics. Although most studies performed on rodents and cell cultures report an upregulation of respective enzymes and transporters, data from the literature are inconsistent. Especially results obtained with sheep differ from these observations. As vitamin D metabolites are widely used as feed additives or therapeutics in livestock animals, we aimed to assess whether these differences indicate species-specific responses or occurred due to the very high dosages used in the rodent studies. Therefore, we applied treatment protocols to rats that had been used previously in sheep or cattle. Forty-eight female rats were divided into three treatment and corresponding placebo groups: (1) a single intraperitoneal injection of 1,25-(OH)2D3 or placebo 12 h before sacrifice; (2) daily supplementation with 25-OHD3 by oral gavage or placebo for 10 days; and (3) a single intramuscular injection of vitamin D3 10 days before sacrifice. In contrast to a previous study using sheep, treatment of rats with 1,25-dihydroxyvitamin D3 did not result in an upregulation of cytochrome P450 3A isoenzymes (CYP3A), but a decrease was found in hepatic and intestinal expressions. In addition, a downregulation of P-glycoprotein (P-gp) and breast cancer resistance protein was found in the brain. Taken together, the stimulating effects of vitamin D metabolites on the expression of genes involved in the metabolism and elimination of xenobiotics reported previously for rodents and sheep could not be reproduced. In contrast, we even observed a negative impact on the expression of CYP3A enzymes and their most important regulator, the pregnane X receptor. Most interestingly, we could demonstrate an effect of treatment with 25-hydroxyvitamin D3 and vitamin D3 on the functional activity of ileal P-glycoprotein (P-gp) using the Ussing chamber technique.

Stealth surface driven accumulation of “Trojan Horse” for tumor hypoxia relief in combination with targeted cancer therapy

The clearance of tumor-associated immune systems (TAISs) is the key impediment of the drug delivery, which leads to the low drug accumulation (0.7%) in the traditional chemotherapy. To tackle this issue, we constructed biomimetic nanoparticles serving as “Trojan Horse”, which consist of polydopamine (PDA) nanospheres with payloads of catalase (CAT), anticarcinogenic drug doxorubicin (Dox) and surrounding tumor cell membrane. Due to the coating of the homotypic cancer cell membrane, the “Trojan Horse” escaped the clearance of TAISs and heightened tumor-site target delivery. Sequentially, the over-expressive hydrogen peroxide in the tumor microenvironment penetrated the membrane of the “Trojan Horse” and reacted with CAT to generate oxygen, which realized the hypoxia relief. The hypoxia relief removed the immune resistance of tumors via the polarization of tumor-associated macrophages from M2 to M1 and broke the chemoresistance by downregulation of P-glycoprotein. With the oxygen generation, the cell membrane was broken and separated from the Dox-loaded PDA resulting in the immunotherapy by antigen presentation. Finally, the Dox-PDA nanospheres were taken into the tumor cells to achieve the chemo-phototherapy under near-infrared laser irradiation. The “Trojan Horse” showed higher accumulation (15.3%) over the enhanced permeability and retention effect (7.5%) of conventional nanodrugs and led to complete tumor eradication in vivo, which provides a promising treatment paradigm against tumor.

The effects of ultrasound exposure on P-glycoprotein-mediated multidrug resistance in vitro and in vivo

BackgroundMultidrug resistance (MDR) is often responsible for the failure of chemotherapy treatment, and current strategies for cancer MDR are not adequately satisfying as to their efficacy and safety. In this study, we sought to determine the anti-MDR effects of ultrasound (US) irradiation and its underlying mechanisms against drug-resistance.MethodsMDR variant MCF-7/ADR cell lines and endothelial cell lines were used to determine the appropriate ultrasound intensity for in vitro experiments. MCF-7/ADR cell and HEPG2/ADM cells were used to assess the anti-MDR effect of US irradiation. Intracellular adriamycin (ADM) accumulation, Cell viability, cell proliferation and cell apoptosis were evaluated after ADM + US treatment or ADM treatment alone. MCF-7/ADR xenograft mice were used to investigate the appropriate ultrasound intensity for in vivo experiments and its effect on the long-term prognosis. Underlining mechanisms by which ultrasound exposure reversing MDR phenotype were investigated both in vitro and in vivo.ResultsCombination of ADM and 0.74 W/cm2 US irradiation enhanced ADM intracellular concentration and nuclear accumulation in MCF-7/ADR and HEPG2/ADM cells, compared to those treated with ADM alone. Enhanced cellular ADM uptake and nuclei localization was associated with increased cytotoxicity of ADM to ADM-resistant cells, lower ADM-resistant cell viability and proliferative cell ratio, and higher apoptotic cell ratio. More importantly, US exposure increased the effectiveness of ADM to inhibit tumor growth in MCF-7/ADR xenograft mice. Mechanistically, US exposure promoted ADM accumulation in MDR cells mainly through down-regulation of P-glycoprotein (P-gp), which is dependent on US-induced intracellular reactive oxygen species (ROS) production. US-induced oxidative stress promoted miR-200c-3p and miR-34a-3p expression by forming miR-200c/34a/ZEB1 double-negative feedback loop. Finally, US-induced miR-200c/34a overexpression decreased P-gp expression and reversed MDR phenotype.ConclusionUS irradiation could reverse MDR phenotype by activating ROS-ZEB1-miR200c/34a-P-gp signal pathway. Our findings offer a new and promising strategy for sensitizing cells to combat MDR and to improve the therapeutic index of chemotherapy.

PPARs are mediators of anti-cancer properties of superparamagnetic iron oxide nanoparticles (SPIONs) functionalized with conjugated linoleic acid

Breast cancer chemotherapy can cause side effects due to nonspecific drug delivery, low solubility and fast metabolism of drugs used in conventional therapy. Moreover, the therapeutic effect of the drugs is often reduced by the strengthening of chemoresistance, which occurs via a variety of mechanisms. Different strategies have been developed to reduce multidrug resistance (MDR)-associated gene expressions including the use of surfactants and polymers. In this study superparamagnetic iron oxide nanoparticles (SPIONs) functionalized with conjugated linoleic acid (CLA) reduced the number and viability of cells in comparison with both untreated cells or cells treated with SPIONs alone. This cytostatic effect correlated with the increase of peroxisome proliferator-activated receptors γ (PPARγ). The necrotic cell death induced, as a consequence, an inflammatory process, as evidenced by the decrease of the anti-inflammatory PPARα and increase of pro-inflammatory TNFα and IL-1β. PPARs were examined because CLA is one of their natural ligands. The antitumor effect observed was accompanied by a down-regulation of p-glycoprotein (P-gp), which was the first important discovered efflux transporter belonging to MDR, and of ALDH3A1, an enzyme able to metabolize some drugs, reducing their effects. The down-regulation of P-gp correlated with the increase of cytokines. The ALDH3A1 decrease correlated with the increase of PPARγ. Based on these results, PPARs are molecular mediators of anti-cancer effect of SPIONs functionalized with CLA, being changes in these nuclear receptors correlated with induction of cytotoxicity and inflammation, and decreased ability of cancer cells in blocking anti-cancer drug effect.

Berberine Upregulates P-Glycoprotein in Human Caco-2 Cells and in an Experimental Model of Colitis in the Rat via Activation of Nrf2-Dependent Mechanisms.

Downregulation of P-glycoprotein (P-gp) is implicated in the pathophysiology of inflammatory bowel disease (IBD). Berberine, a principal isoquinoline alkaloid extracted from Berberis species, has been reported to exhibit therapeutic potential in IBD. In this study, we used a dextran sulfate sodium (DSS)-induced colitis rat model to evaluate the effect of berberine on P-gp and explore its mechanism of action. Berberine treatment improved DSS-induced colitis symptoms, attenuated inflammatory markers (myeloperoxidase, tumor necrosis factor-α, and interleukin-1β and -6), and enhanced P-gp expression in a dose-dependent manner. Although colonic expression of the P-gp-related nuclear receptor pregnane X receptor and transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) were downregulated in the colitis model, gene and protein expression analysis revealed that berberine treatment reversed only the downregulation of Nrf2. In vitro studies using Caco-2 cells showed that the multidrug resistance 1 (MDR1) gene and P-gp protein were upregulated by berberine in a dose- and time-dependent manner. Significant upregulation of the MDR1 gene by berberine was abrogated by Nrf2 silencing, indicating that the Nrf2-mediated pathway was responsible for this activation. Luciferase assays showed a dose-dependent increase in Nrf2 reporter gene activity after berberine treatment in Caco-2 cells, with a significant 2-fold elevation at 2.5 μM berberine, suggesting that berberine is a strong Nrf2 activator. These results indicate the possible involvement of Nrf2-mediated upregulation of P-gp in the therapeutic effect of berberine on colitis and highlight the potential of P-gp and/or Nrf2 as new therapeutic targets for IBD.

Paris saponin VII induces cell cycle arrest and apoptosis by regulating Akt/MAPK pathway and inhibition of P-glycoprotein in K562/ADR cells.

Paris saponinVII (PSVII) is a steroidal saponin isolated from the roots and rhizomes of Trillium tschonoskii Maxim. We found that PSVII could inhibit the growth of adriamycin-resistant human leukemia cells (K562/ADR) in a dose-dependent manner. Furthermore, the molecular mechanism underlying the cytotoxicity and downregulation of P-glycoprotein (P-gp) expression by PSVII was clarified. PSVII significantly suppressed cell proliferation by cell cycle arrest in the G0/G1 phase, which was associated with an obvious decrease in cyclin B1/D1 and CDK2/4/6 protein expression. Moreover, PSVII could attenuate mitochondrial membrane potential, increase the expression of apoptosis-related proteins, such as Bax and cytochrome c, and decrease the protein expression levels of Bcl-2, caspase-9, caspase-3, PARP-1, and p-Akt. We also found that JNK, ERK1/2, and p38 were regulated by PSVII in K562/ADR cells. And further studies indicated that the decrease in the reactive oxygen species level inhibited intrinsic P-gp expression. Therefore, PSVII-induced apoptosis in K562/ADR cells was associated with Akt/MAPK and the inhibition of P-gp. In addition, PSVII induced a robust autophagy in K562/ADR cells as demonstrated by the degradation of LC3-I. These results provide a biochemical basis for possible clinical applications of PSVII in the treatment of leukemia.

Emodin reverses leukemia multidrug resistance by competitive inhibition and downregulation of P-glycoprotein.

Development of multidrug resistance (MDR) is a continuous clinical challenge partially due to the overexpression of P-glycoprotein (P-gp) for chronic myelogenous leukemia (CML) patients. Herein, we evaluated the inhibitory potency of emodin, a natural anthraquinone derivative isolated from Rheum palmatum L, on P-gp in P-gp positive K562/ADM cells. Competition experiments combined with molecular docking analysis were utilized to investigate the binding modes between emodin and binding sites of P-gp. Emodin reversed adriamycin resistance in K562/ADM cells accompanied with the decrease of P-gp protein expression, further increasing the uptake of rhodamine123 in both K562/ADM and Caco-2 cells, indicating the inhibition of P-gp efflux function. Moreover, when incubated with emodin under different conditions where P-gp was inhibited, K562/ADM cells displayed increasing intracellular uptake of emodin, suggesting that emodin may be the potential substrate of P-gp. Importantly, rhodamine 123 could increase the Kintrinsic (Ki) value of emodin linearly, whereas, verapamil could not, implying that emodin competitively bound to the R site of P-gp and noncompetition existed between emodin and verapamil at the M site, in a good accordance with the results of molecular docking that emodin bound to the R site of P-gp with higher affinity. Based on our results, we suggest that emodin might be used to modulate P-gp function and expression.

Multifunctional nanoparticles combining host-guest interaction of CB[7] and AuNS for light-promoted endosome escape and high gene transfection

Breast cancer chemotherapy can cause side effects due to nonspecific drug delivery, low solubility and fast metabolism of drugs used in conventional therapy. Moreover, the therapeutic effect of the drugs is often reduced by the strengthening of chemoresistance, which occurs via a variety of mechanisms. Different strategies have been developed to reduce multidrug resistance (MDR)-associated gene expressions including the use of surfactants and polymers. In this study superparamagnetic iron oxide nanoparticles (SPIONs) functionalized with conjugated linoleic acid (CLA) reduced the number and viability of cells in comparison with both untreated cells or cells treated with SPIONs alone. This cytostatic effect correlated with the increase of peroxisome proliferator-activated receptors γ (PPARγ). The necrotic cell death induced, as a consequence, an inflammatory process, as evidenced by the decrease of the anti-inflammatory PPARα and increase of pro-inflammatory TNFα and IL-1β. PPARs were examined because CLA is one of their natural ligands. The antitumor effect observed was accompanied by a down-regulation of p-glycoprotein (P-gp), which was the first important discovered efflux transporter belonging to MDR, and of ALDH3A1, an enzyme able to metabolize some drugs, reducing their effects. The down-regulation of P-gp correlated with the increase of cytokines. The ALDH3A1 decrease correlated with the increase of PPARγ. Based on these results, PPARs are molecular mediators of anti-cancer effect of SPIONs functionalized with CLA, being changes in these nuclear receptors correlated with induction of cytotoxicity and inflammation, and decreased ability of cancer cells in blocking anti-cancer drug effect.

Pharmacokinetics of talinolol is modified by barnidipine: implication of P-glycoprotein modulation.

Concomitant administration of P-glycoprotein substrates and inhibitors may cause pharmacokinetic drug interactions leading to increased concentrations associated with serious side effects and toxicities. Barnidipine is a longacting calcium-channel blocker and potent inhibitor of P-glycoprotein in vitro, and talinolol is a beta-blocker and probe substrate of P-glycoprotein. This study was designed to investigate the effects of single and repeated oral doses of barnidipine on talinolol pharmacokinetics in rats. In the single-dose study, talinolol (20 mg/kg) alone and with barnidipine at low (1 mg/kg) and high doses (10 mg/kg) were orally administered to rats. In the repeated-dose study, rats were treated with barnidipine (1 mg/kg/day) or vehicle only for four days, then with talinolol (20 mg/kg, on day 5). Blood samples were collected at 0.5, 1, 2, 4, 6 h following last dose and plasma talinolol levels were determined by HPLC. Compared to the control, Cmax of talinolol elevated 10% (p=0.79) and 110% (p<0.05); plasma AUC0-6h increased 33% (p=0.41) and 46% (p<0.05) following low and high single doses of barnidipine co-administration, respectively. In the repeated-dose study, Cmax and AUC0-6h of talinolol increased 131% (p<0.05) and 130% (p<0.05) respectively, following co-administration of a low barnidipine dose. Double-peaks were observed when single or repeated low doses of barnidipine were co-administered. There may be coupling between occurrence of double-peak phenomenon and P-glycoprotein inhibition. Increment of talinolol bioavailability upon low and high doses of barnidipine co-administration may be due to P-glycoprotein inhibition. The higher increase of talinolol plasma AUC0-6h due to the repeated doses of barnidipine may be explained by downregulation of P-glycoprotein.