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Wu CP, Calcagno AM, Hladky SB, Ambudkar SV, Barrand MAModulatory effects of plant phenols on human multidrug-resistance proteins 1, 4 and 5 (ABCC1, 4 and 5). FEBS J 272: 4725-4740
The FEBS Journal
Abstract
Plant flavonoids are polyphenolic compounds, commonly found in vegetables, fruits and many food sources that form a significant portion of our diet. These compounds have been shown to interact with several ATP-binding cassette transporters that are linked with anticancer and antiviral drug resistance and, as such, may be beneficial in modulating drug resistance. This study investigates the interactions of six common polyphenols; quercetin, silymarin, resveratrol, naringenin, daidzein and hesperetin with the multidrug-resistance-associated proteins, MRP1, MRP4 and MRP5. At nontoxic concentrations, several of the polyphenols were able to modulate MRP1-, MRP4- and MRP5-mediated drug resistance, though to varying extents. The polyphenols also reversed resistance to NSC251820, a compound that appears to be a good substrate for MRP4, as predicted by data-mining studies. Furthermore, most of the polyphenols showed direct inhibition of MRP1-mediated [3H]dinitrophenyl S-glutathione and MRP4-mediated [3H]cGMP transport in inside-out vesicles prepared from human erythrocytes. Also, both quercetin and silymarin were found to inhibit MRP1-, MRP4- and MRP5-mediated transport from intact cells with high affinity. They also had significant effects on the ATPase activity of MRP1 and MRP4 without having any effect on [32P]8-azidoATP[alphaP] binding to these proteins. This suggests that these flavonoids most likely interact at the transporter's substrate-binding sites. Collectively, these results suggest that dietary flavonoids such as quercetin and silymarin can modulate transport activities of MRP1, -4 and -5. Such interactions could influence bioavailability of anticancer and antiviral drugs in vivo and thus, should be considered for increasing efficacy in drug therapies.
... Notably, ABCG1 has already been demonstrated in vitro (in macrophages) to be induced by QUE, thus protecting against ox-LDL-induced injury [106]. Looking at the xenobiotic efflux transporters, which are known to interact with several polyphenols [107], the role of ABCB1 in the transport of AFB1 is still unclear. Indeed, contrasting results were reported in vitro and in vivo [108,109], preventing us from interpreting the significance of ABCB1 downregulation observed here in the co-treatment with QUE and AFB1. ...
... On the other hand, QUE has been reported to negatively affect ABCB1 activity in HepG2 cells, thus potentiating the cytotoxic activity of anticancer drugs [112]. With regard to MRP4, it has been shown that QUE is most probably a substrate for this transporter, and its activity can be modulated by flavonoids [107]. To our knowledge, there is no evidence of a possible relationship between AFB1 and MRP4; however, the expression of ABCC4 is upregulated in HCC tissues. ...
Aflatoxin B1 (AFB1) induces lipid peroxidation and mortality in bovine foetal hepatocyte-derived cells (BFH12), with underlying transcriptional perturbations associated mainly with cancer, cellular damage, inflammation, bioactivation, and detoxification pathways. In this cell line, curcumin and resveratrol have proven to be effective in mitigating AFB1-induced toxicity. In this paper, we preliminarily assessed the potential anti-AFB1 activity of a natural polyphenol, quercetin (QUE), in BFH12 cells. To this end, we primarily measured QUE cytotoxicity using a WST-1 reagent. Then, we pre-treated the cells with QUE and exposed them to AFB1. The protective role of QUE was evaluated by measuring cytotoxicity, transcriptional changes (RNA-sequencing), lipid peroxidation (malondialdehyde production), and targeted post-transcriptional modifications (NQO1 and CYP3A enzymatic activity). The results demonstrated that QUE, like curcumin and resveratrol, reduced AFB1-induced cytotoxicity and lipid peroxidation and caused larger transcriptional variations than AFB1 alone. Most of the differentially expressed genes were involved in lipid homeostasis, inflammatory and immune processes, and carcinogenesis. As for enzymatic activities, QUE significantly reverted CYP3A variations induced by AFB1, but not those of NQO1. This study provides new knowledge about key molecular mechanisms involved in QUE-mediated protection against AFB1 toxicity and encourages in vivo studies to assess QUE’s bioavailability and beneficial effects on aflatoxicosis.
... The main flavonoids present in C. sinensis extract include catechins (flavan-3-ols) such as epigallocatechin-3gallate, epigallocatechin, epicatechin-3-gallate, and epicatechin; it also contains gallic acid, chlorogenic acid, caffeic acid, and flavonol derivatives such as kaempferol, myricetin, and quercetin, which are other constituents of green tea. 35,36 The chemical structure of the polyphenols allows them to cross through the blood-brain barrier, [37][38][39][40][41][42][43][44][45][46][47] and its intake has been associated with neuronal protection against toxicant agents' impact. Green tea research shows an amelioration of behavioral aberrations by a daily intake dose of the extract; doses range from 75 to 300 mg/kg after postnatal day 14 up to day 40 in animals with valproate induced autism. ...
... 53 In recent years, ASD cases have been increasing from 1/20 live births in 2006 54 up to 1/68 live births in 2014 55 ; nonetiological causes have been determined yet, which hampered the development of treatments that contributes to alleviate ASD symptoms. Nowadays, phytopharmaceutical treatments have been tried out in clinical trials; one of the flavones that shows anti-inflammatory, antioxidant, antiallergy, and neuroprotective effects is Luteolin (NeuroProtek Ò ) with a mixture of the flavonoids Quercetin and Rutin, which are structurally related in a liposomal formulation, 56 these compounds have potent inhibition of histamine, interleukin (IL)-6 from microglia, 44 IL-8, TNF, and tryptase release from human mast cells, 57 and they also block mercury-induced cytokine release from human mast cells. 58,59 Luteolin formulation (NeuroProtek) research in children with ASD states the improvement of ASD symptoms in those who used the formula for at least 4 months; 75% (28/37) of patients reported significant improvement in features such as their stool shape, smell, form, color, and habits within a period of 2 to 3 weeks. ...
Over the years, scientific researches have validated the healing benefits of many psychopharmacotherapeutic plant-based drugs to ameliorate psychiatric disorders. In contrast, the use of chemical procedures to isolate and purify specific compounds from plants that have been used to treat autism spectrum disorders (ASDs) and its clinical features may contribute to improve the quality of life of many patients. Also, herbal pharmacological treatments could improve the core symptoms of autism with fewer side effects. This review will focus on the uses and actions of phytopharmaceuticals in the behavioral conditions of ASDs. A large number of natural compound-based plant drugs have been tested in murine models of autism and in clinical trials with remarkable success in reversing the core and associated behaviors with autism such as flavonoids, cannabinoids, curcuminoids, piperine, resveratrol, and bacosides. This plant-based drug alternative is safer given that many psychiatric disorders and neurodegenerative pathologies do not often respond well to currently prescribed medications or have significant side effects. However, it is noteworthy to consider the need for large clinical trials to determine safety and efficacy. Many results are based on case reports or small size samples, and often the studies are open label. Standardization of procedures (i.e., purity and concentrations) and quality controls are strictly required to ensure the absence of side effects.
... In the previous two sections, many substrates of MRP1 have already been described, such as LTC 4 , 26,27,29 64,88,136,239,252,258,316 or GSH. 23-29,32,43,45,86,94,98,101,122-124,133,134,183,189,205,206,237,248,252,253,258,265,279,297,312,320-322 324-326,352 Besides, some inhibitors got attention since they have been widely used as standard inhibitors of this transporter, such as MK571, 24,44,93,133,134,151,179,183,189,202,247,251,252,273,276,290,291,352 or BSO. 43,44,122,133,134,189,352 This section focuses on both the already mentioned inhibitors (in more detail), and others that are either derived from intrinsic substrates of MRP1 or related compounds. ...
... The authors showed that resveratrol downregulated MRP1 on mRNA as well as on protein level at concentrations between 50 and 100 M within 24 hr. Additionally, this compound was shown by Wu et al. to stimulate MRP1 ATPase by 49% at a concentration of 30 M.247 Piceatannol, already described above as a rather weak MRP1 inhibitor,93 was found by Wesolowska et al.to reduce BCECF export in erythrocytes by 61.3%.246 Additionally, the authors reported that the stilbene derivatives trans-3,5,3 ′ ,4 ′ -tetraacetoxystilbene and trans-3,5,3 ′ ,4 ′ -tetramethoxystilbene (Figure 23) were inhibitors of MRP1, which reduced BCECF transport by 56.3% and 28.5%, respectively, at 75 M. ...
Multidrug resistance‐associated protein 1 (MRP1, ABCC1) is an ATP‐binding cassette (ABC) transport protein. This efflux pump uses the energy of ATP hydrolysis to export structurally diverse antineoplastic agents in human cancers. The upregulation of MRP1 (either inherent or acquired) is one major reason for the occurrence of the phenomenon called multidrug resistance (MDR). MDR is characterized by a reduced outcome of chemotherapy due to the active intracellular clearance of cytostatic drugs below the necessary effect concentration. Much effort has been made to overcome MDR, which implied high‐throughput screenings of already known pharmacological and natural compounds, modification of intrinsic substrates, as well as design and synthesis of new inhibitors. This review is meant not only to summarize the most recent results over the past 10 years, but also to highlight major achievements regarding reversal of MRP1‐mediated MDR, from the time of its discovery until today. The focus lies on small‐molecule compounds that feature either direct MRP1 inhibition/transport blockage, toxicity against MRP1‐overexpressing cells, inhibition/modification of intracellular processes necessary for MRP1 function, or modification of MRP1‐related metabolic and genomic mechanisms. Considering all aspects, this review might be useful to (re)consider possible strategies to overcome MRP1‐mediated MDR. Furthermore, it may be the basis for developing new, even better, highly potent, less toxic, and selective (as well as broad‐spectrum) MRP1 inhibitors that will enter clinical evaluations in different malignancies and finally conduce to overcome MDR in general.
... In comparison to the antibacterial action of onion, hardly any work has been done to investigate its antiviral properties. In addition to sulphur compounds, it has been reported that quercetin (3,5,7,3',4'tetrahydroxyflavone), the major onion flavonoid, also possesses antiviral activity and enhances the bioavailability of some antiviral drugs (Wu et al., 2005). Onion lectins, unlike the garlic lectins, have a pronounced anti-HIV activity (Van Damme et al., 1993). ...
... Particularly, quercetin and derivatives exhibit anticancer properties, which have been demonstrated in a number of malignancies, including prostate, breast, skin, lung and liver cancers (Avila et al., 1994;Musonda and Chipman, 1998;Le Marchand et al., 2000;Le Marchand, 2002;Vijayababu et al., 2006;Arung, Furuta, Ishikawa, Kusuma, Shimizu, and Kondo, 2011). Likewise, several studies have reported that quercetin enhances bioavailability of some anticancer drugs, as Tamoxifen, a non-steroidal antiestrogen for the treatment and prevention of breast cancer, by promoting their intestinal absorption and reducing their metabolism (Shin et al., 2006;Wu, et al, 2005). ...
Onion (Allium cepa L.) is an important vegetable traditionally used as a food ingredient in the Mediterranean diet that has a high production, domestic, and foreign trade worldwide. It is consumed raw, cooked or processed into different onion products in the daily diet. Onion added into different foods makes these products rich in bioactive compounds with potential beneficial health effects. Among them, its effect on cardiovascular disease, including hypocholesterolemic, hypolipidemic, anti-hypertensive, antithrombotic, and hypoglycaemic activities, is one of the most extensively studied benefits. Onion consumption has also been reported to have antiproliferative effects in many cancer cell lines, to be involved in the bone metabolism and in the behaviour as a possible antidepressant agent, and to stimulate the growth of specific microorganisms in the colon (Bifidobacteria and Lactobacilli) with a general positive health effect. Moreover, traditionally, in the folk medicine, it has been described the use of onion as an antimicrobial, antioxidant, antiinflamatory and asthma-protective agent.
Evidence from several investigations suggests that these biological and medical functions are mainly due to the high organo-sulphur compounds content of onion. Along with organo-sulphur compounds, flavonols (quercetin and its glucosides) and dietary fibre (fructans and fructooligosaccharides (FOS)) have been also related to the onion biological properties. Moreover, recently, it has been demonstrated that additional onion constituents such as saponins and peptides have potentially beneficial health effects, including antifungal, antitumor, antispasmodic and cholesterol-lowering activities and capacity to inhibit the development and activity of osteoclasts in vitro.
As with every biologically active substance, with onion and its derivatives it is necessary to consider certain precautions to minimize the risk of adverse side effects. However, the usefulness of onion as therapeutic agent seems to be very safe, since all its possible adverse effects, such as gastrointestinal upsets and dermatological problems appear with an excessive and prolonged consumption.
... Similar studies have shown that phenolic compounds have no significant effect on cell viability in HEK293 cells. In agreement with these results, Wu et al. [38] reported that phenolic compounds showed no significant (P < .05) effect on human cell HEK293 viability but did enhance the sensitivity of MRP1-HEK293 cells to topside. ...
This study attempted to optimize the extraction conditions (liquid‒solid ratio, temperature and time) of phenolic yield from Cleome (Cleome arabica L.) fruits extract by the surface response methodology “Box‒Behnken plan” to identify and quantify polyphenolic compounds and test their antioxidant activity. A molecular docking method was developed to clarify the anti-apoptotic effects of bioactive molecules of cleome fruits. Optimal conditions were found at a ratio of 8 ml/g, 4°C and 2 h when the maximum yield was 27.24%. The total phenolic and flavonoid content assessed by the Folin-Ciocalteu and Quercetin methods were 195.20 mg gallic acid equivalent/g dry weight (mg GAE/g DW) and 51.20 mg quercetin equivalent/g dry plant weight (mg QE/g DW), respectively. HPLC and FT-IR protocols revealed the presence of five polyphenolic compounds of which catechin was found to be the most abundant. The antioxidant activity of the extract was quantified by DPPH and FRAP tests, and the IC50 values reached 6.125 mg/ml and 2.9 mg/ml, respectively. In silico analysis confirmed the beneficial effects of phenolic compounds. The results revealed significant antioxidant and antiapoptotic activities of phenolic compounds of cleome fruits and may be useful in the pharmaceutical and food industries with appreciable human health-promoting properties.
... Quercetin also blocks the entry of the viral receptor complex into the cell and interrupts its life cycle, leading to viral death (64). In addition, as an essential consideration in developing new antiviral drugs (65,66), quercetin can increase the bioavailability of antiviral drugs in the body. Luteolin has potent anti-inflammatory, antiviral, antibacterial, and other effects (67,68). ...
The global epidemic has been controlled to some extent, while sporadic outbreaks still occur in some places. It is essential to summarize the successful experience and promote the development of new drugs. This study aimed to explore the common mechanism of action of the four Chinese patent medicine (CPMs) recommended in the Medical Observation Period COVID-19 Diagnostic and Treatment Protocol and to accelerate the new drug development process. Firstly, the active ingredients and targets of the four CPMs were obtained by the Chinese medicine composition database (TCMSP, TCMID) and related literature, and the common action targets of the four TCMs were sorted out. Secondly, the targets of COVID-19 were obtained through the gene-disease database (GeneCards, NCBI). Then the Venn diagram was used to intersect the common drug targets with the disease targets. And GO and KEGG pathway functional enrichment analysis was performed on the intersected targets with the help of the R package. Finally, the results were further validated by molecular docking and molecular dynamics analysis. As a result, a total of 101 common active ingredients and 21 key active ingredients of four CPMs were obtained, including quercetin, luteolin, acacetin, kaempferol, baicalein, naringenin, artemisinin, aloe-emodin, which might be medicinal substances for the treatment of COVID-19. TNF, IL6, IL1B, CXCL8, CCL2, IL2, IL4, ICAM1, IFNG, and IL10 has been predicted as key targets. 397 GO biological functions and 166 KEGG signaling pathways were obtained. The former was mainly enriched in regulating apoptosis, inflammatory response, and T cell activation. The latter, with 92 entries related to COVID-19, was mainly enriched to signaling pathways such as Coronavirus disease—COVID-19, Cytokine-cytokine receptor interaction, IL-17 signaling pathway, and Toll-like receptor signaling pathway. Molecular docking results showed that 19/21 of key active ingredients exhibited strong binding activity to recognized COVID-19-related targets (3CL of SARS-CoV-2, ACE2, and S protein), even better than one of these four antiviral drugs. Among them, shinflavanone had better affinity to 3CL, ACE2, and S protein of SARS-CoV-2 than these four antiviral drugs. In summary, the four CPMs may play a role in the treatment of COVID-19 by binding flavonoids such as quercetin, luteolin, and acacetin to target proteins such as ACE2, 3CLpro, and S protein and acting on TNF, IL6, IL1B, CXCL8, and other targets to participate in broad-spectrum antiviral, immunomodulatory and inflammatory responses.
... People worldwide use it as a spice to improve the taste and smell of food. Also, onions have medicational features such as anti-cancer (Block, 1985), antimicrobial (Griffiths et al., 2002), antiviral (Wu et al., 2005), antifungal (Lun et al., 1994), and also extracts and essential oils of these plants are effective in treating cardiovascular diseases (Rahman & Lowe, 2006). One of the primary health concerns in Iran is food safety as reports of increasing contamination of food with heavy metals are increasing (Fakhri et al., 2018b;Shokri et al., 2021). ...
Considering the importance of onions consumption in the household diet, the control of heavy elements' concentration in foodstuffs is important to ensure the safety of an individual's health. This study aimed to evaluate the risk of heavy metals through onion consumption on human health. In this cross-sectional experimental study, 22 onion samples with varieties red, yellow, and white in the two seasons of autumn and winter in 2020 were randomly collected from the different provinces of Kurdistan, Hamedan, and Kermanshah. The concentrations of heavy metals were evaluated with an atomic absorption spectrometer. The risks of human health were evaluated by the hazard quotient (HQ) and the obtained results were analyzed with one-way ANOVA and t-test. The obtained findings demonstrated that all collected samples contained heavy metals. For example, the cadmium (Cd) concentration in onion samples in the province of West Azerbaijan, Kurdistan, Hormozgan, Isfahan, and Zanjan was 526.49, 274.49, 69.77, 67.39, 65.69 µg kg⁻¹, respectively. While the standard specified in Iran for the concentration of Cd in onions is 50 μg kg⁻¹. However, the rate of lead (Pb) contamination in samples collected from Isfahan, Hormozgan, Zanjan Khuzestan, Tehran (Varamin) was 296.50, 266.71, 261.49, 215.64, 106.19 µg kg⁻¹, respectively, which less than maximum allowable limit recommended by WHO-FAO (300 µg kg⁻¹). The HQ for non-cancerous diseases for Cd and Pb were 8.6× 10⁻² and 1.6× 10⁻¹, respectively, and the probability of carcinogenic risk for Pb (8.1× 10⁻⁴) was at the level of acceptable. There is no concern about the non-carcinogenic diseases and carcinogenic risk of consuming heavy metals in onion. Therefore, for optimal management and prevention of further pollution, it is recommended to study the origin and determine the amounts of heavy metals for their potential contamination of foodstuffs from the soil, water, and dust in the region.
... Aromatase inhibition, anti-inflammatory mechanisms, antioxidant mechanisms, and anti-estrogenic processes can all be attributed to the polyphenol's powerful anticancer effect [29][30][31][32][33][34]. Aromatase is basically an estrogen synthase that is basically expressed in the tissue of breast cancer. Flavones such as luteolin and isoflavone have been shown to bind active sites of aromatase as estrogen receptors [35]. In carcinogenesis, an imbalance in the actions of cell cycle advancement proteins (Cyclins and cyclin-dependent kinases or CDK) and cell cycle arrest proteins (CDK inhibitors) causes significant cell proliferation. ...
The given review summarizes the various molecular mechanisms responsible for the metabolic action of flavonoids as anticancer agents. Various types of flavonoids have proven to show biological & pharmacological activities like anti-inflammatory, antimicrobial, antioxidant, anticancer, anti-allergic, and antidiarrheal activities. The chemoprotective nature of flavonoids is also discussed due to their ability to inhibit topoisomerase enzymes at various stages of cancer, such as tumor initiation, promotion, and progression. The various biomolecular activities which are responsible for their role as the chemopreventive agent may be due to their antioxidative effect, anti-angiogenic properties, induction of protective enzymes, inhibitory action on the cell-like protein kinase activity inhibition, spreading of tumor cells, apoptosis induction, tumor cell invasion to name a few. There has been much-emerging evidence based on the versatility of flavonoids, their complex mechanism of action, lesser side effects, and varied pharmacological properties that make them potential anticancer agents. Challenges associated with their use in extraction, isolation, purification, and checking bioefficacy are also discussed.
... They are known to have many prominent health benefits [712,713], including anticancer properties [714][715][716]. In terms of MDR, flavonoids have been studied and characterized extensively by many research groups to determine their capability to restore drug sensitivity in MDR tumor cells [717][718][719][720]. ...
Cancer is one of the leading causes of death globally. The development of drug resistance is the main contributor to cancer-related mortality. Cancer cells exploit multiple mechanisms to reduce the therapeutic effects of anticancer drugs, thereby causing chemotherapy failure. Natural products are accessible, inexpensive, and less toxic sources of chemotherapeutic agents. Additionally, they have multiple mechanisms of action to inhibit various targets involved in the development of drug resistance. In this review, we have summarized the basic research and clinical applications of natural products as possible inhibitors for drug resistance in cancer. The molecular targets and the mechanisms of action of each natural product are also explained. Diverse drug resistance biomarkers were sensitive to natural products. P-glycoprotein and breast cancer resistance protein can be targeted by a large number of natural products. On the other hand, protein kinase C and topoisomerases were less sensitive to most of the studied natural products. The studies discussed in this review will provide a solid ground for scientists to explore the possible use of natural products in combination anticancer therapies to overcome drug resistance by targeting multiple drug resistance mechanisms.
... Substrates of MRP4 include cyclic adenosine monophosphate (cAMP), cyclic guanosine monophosphate (cGMP), adefovir, ganciclovir, loop diuretics, cephalosporins, topotecan, PMEA, 6-mercaptopurine (6-MP), 6-thioguanine (6-TG), etc. [210][211][212]214]. Recent updates added a few more natural products like plant polyphenols, resveratrol, and quercetin that are transported by MRP4 [213]. ...
The phenomenon of drug resistance has been a hindrance to therapeutic medicine since the late 1940s. There is a plethora of factors and mechanisms contributing to progression of drug resistance. From prokaryotes to complex cancers, drug resistance is a prevailing issue in clinical medicine. Although there are numerous factors causing and influencing the phenomenon of drug resistance, cellular transporters contribute to a noticeable majority. Efflux transporters form a huge family of proteins and are found in a vast number of species spanning from prokaryotes to complex organisms such as humans. During the last couple of decades, various approaches in analyses of biochemistry and pharmacology of transporters have led us to understand much more about drug resistance. In this review, we have discussed the structure, function, potential causes, and mechanisms of multidrug resistance in bacteria as well as cancers.
... An improved formulation of quercetin for oral absorption has been recently described from a clinical study 46 . Additionally, quercetin at concentration up to 10 µM inhibits the ATPase of proteins associated with multi-drug resistance 47 . This may support the applicability of quercetin for pharmaceutical purposes. ...
O’nyong-nyong virus is an alphavirus closely related to chikungunya virus, causing arthralgia, rash and fever. Alphaviruses mainly target synovial fibroblasts and persists in the joints of patients, possibly leading to chronic arthritis. To date, no specific antiviral treatment is available for ONNV infection and induced-inflammation. Primary human synovial fibroblasts cells were used to assess infection by ONNV and the resulting cytokine responses. Phenolics (gallic acid, caffeic acid and chlorogenic acid, curcumin and quercetin) and a curcuminoids-rich extract from turmeric were tested for their antiviral and anti-inflammatory capacities. We showed that infection occurred in HSF cells and increased gene expression and protein secretion of two major proinflammatory CCL-2 and IL-1β markers. In ONNV-infected HSF cells (MOI 1), we found that non-cytotoxic concentrations of phenolics (10 µM) reduced the level of viral RNA (E1, E2, nsP1, nsP2) and downregulated CCL-2 and IL-1β expression and secretion. These results highlighted the high value of the flavonol quercetin to reduce viral RNA levels and inflammatory status induced by ONNV in HSF cells.
... Quercetin is report to modulate the transport activities of the multidrug-resistance-associated proteins. Moreover, quercetin could not only influence the bioavailability of anticancer and antiviral drugs in vivo but also alleviate endotoxin-stimulated apoptosis and inflammation by up-regulation of relevant micro-RNA [41,42]. Additionally, quercetin could mitigate the intracellular formation of reactive oxygen species (ROS) to reduce the pro-inflammatory response [43]. ...
Introduction
Shuanghuanglian (SHL) oral liquid is a well-known traditional Chinese medicine preparation administered for respiratory tract infections in China. However, the underlying pharmacological mechanisms remain unclear. The present study aims to determine the potential pharmacological mechanisms of SHL oral liquid based on network pharmacology.
Methods
Network pharmacology-based strategy including collection and analysis of putative compounds and target genes, network construction, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, and Gene Ontology(GO) enrichment, identification of key compounds and target genes, and molecule docking was performed in this study.
Results
A total of 82 bioactive compounds and 226 putative target genes of SHL oral liquid were collected. Of note, 28 hub target genes including 4 major hub target genes: estrogen receptor 1 (ESR1), nuclear receptor coactivator 2 (NCOA2), nuclear receptor coactivator 1 (NCOA1), androgen receptor (AR) and 5 key compounds (quercetin, luteolin, baicalein, kaempferol and wogonin) were identified based on network analysis. The hub target genes mainly enriched in pathways including PI3K-Akt signaling pathway, human cytomegalovirus infection, and human papillomavirus infection, which could be the underlying pharmacological mechanisms of SHL oral liquid for treating diseases. Moreover, the key compounds had great molecule docking binding affinity with the major hub target genes.
Conclusion
Using network pharmacology analysis, SHL oral liquid was found to contain anti-virus, anti-inflammatory, and “multi-compounds and multi-targets” with therapeutic actions. These findings may provide a valuable direction for further clinical application and research.
... Onion bulbs (Allium cepa) encompass a myriad of bioactive natural product classes which are linked to its health benefits, i.e., anticancer or chemo-preventive agents [17]; antimicrobial [18], antiviral [19], and antioxidant activities [20]; protective activities against cardiovascular diseases [21], diabetes [22], neurodegenerative disorders, and cataract formation [23,24]. Among the secondary metabolites found in onion, flavonoids, fructans, and organosulphur compounds were mainly found to mediate its health benefits [25][26][27]. ...
Brine, the historically known food additive salt solution, has been widely used as a pickling media to preserve flavor or enhance food aroma, appearance, or other qualities. The influence of pickling, using brine, on the aroma compounds and the primary and secondary metabolite profile in onion bulb Allium cepa red cv. and lemon fruit Citrus limon was evaluated using multiplex metabolomics technologies. In lemon, pickling negatively affected its key odor compound "citral", whereas monoterpene hydrocarbons limonene and γ-terpinene increased in the pickled product. Meanwhile, in onion sulphur rearrangement products appeared upon storage, i.e., 3,5-diethyl-1,2,4-trithiolane. Profiling of the polar secondary metabolites in lemon fruit via ultra-performance liquid chromatography coupled to MS annotated 37 metabolites including 18 flavonoids, nine coumarins, five limonoids, and two organic acids. With regard to pickling impact, notable and clear separation among specimens was observed with an orthogonal projections to least squares-discriminant analysis (OPLS-DA) score plot for the lemon fruit model showing an enrichment of limonoids and organic acids and that for fresh onion bulb showing an abundance of flavonols and saponins. In general, the pickling process appeared to negatively impact the abundance of secondary metabolites in both onion and lemon, suggesting a decrease in their food health benefits.
... As well as this, cell culture data have shown that the quercetin flavonol may inhibit the replication of different respiratory viruses, reducing their viral count (Chiang et al., 2003;Chen et al., 2006). Quercetin also enhances the bioavailability of some anti viral drugs (Wu et al., 2005). Lectins are a very heteroge neous group of glycoproteins, with the ability to recognize and bind specifically to carbohydrate ligands. ...
... Although phloretin is used as an AQP9 inhibitor it is also known to inhibit numerous other unrelated transport proteins (Krupka, 1985;Kwon et al., 2007;Shirasaka et al., 2013;Wang and Morris, 2007). Phloretin is a bioflavonoid and compounds from this class can be potent inhibitors of MRPs (Wu et al., 2005). The influence of phloretin on MRP4 activity was unknown, thus the effect of increasing concentrations of phloretin on E 2 17bG transport by membrane vesicles prepared from HEK293 cells over-expressing MRP4 was evaluated. ...
Arsenic is a proven human carcinogen and associated with a myriad of other adverse health effects. This metalloid is methylated in human liver to monomethylarsonic acid (MMA(V)), monomethylarsonous acid (MMA(III)), dimethylarsinic acid (DMA(V)), and dimethylarsinous acid (DMA(III)) and eliminated predominantly in urine. Hepatic basolateral transport of arsenic species is ultimately critical for urinary elimination; however, these pathways are not fully elucidated in humans. A potentially important human hepatic basolateral transporter is the ATP-binding cassette (ABC) transporter multidrug resistance protein 4 (MRP4/ABCC4) that in vitro is a high affinity transporter of DMA(V) and the diglutathione conjugate of MMA(III) [MMA(GS)2]. In rats, the related canalicular Mrp2/Abcc2 is required for biliary excretion of arsenic as As(GS)3 and MMA(GS)2. The current study used sandwich cultured human hepatocytes (SCHH) as a physiological model of human arsenic hepatobiliary transport. Arsenic efflux was detected only across the basolateral membrane for nine out of fourteen SCHH preparations, five had both basolateral and canalicular efflux. Basolateral transport of arsenic was temperature- and GSH-dependent and inhibited by the MRP inhibitor MK-571. Canalicular efflux was completely lost after GSH depletion suggesting MRP2-dependence. Treatment of SCHH with As(III) (0.1-1 μM) dose-dependently increased MRP2 and MRP4 levels, but not MRP1, MRP6, or aquaglyceroporin 9. Treatment of SCHH with oltipraz (Nrf2 activator) increased MRP4 levels and basolateral efflux of arsenic. In contrast, oltipraz increased MRP2 levels without increasing biliary excretion. These results suggest arsenic basolateral transport prevails over biliary excretion and is mediated at least in part by MRPs, most likely including MRP4.
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... As reported previously, the MRP1-overexpressing cell lines, HEK293/MRP1 and HL60/Adr, showed high resistance to vincristine, vinblastine, etoposide and doxorubicin (Wu et al., 2005). In the present research, we showed that ibrutinib, at concentrations showing little or no toxicity, significantly potentiated the cytotoxicity of known MRP1 substrates in MRP1-overexpressing cells. ...
Background and purpose:
The transporter, multidrug resistance protein 1 (MRP1, ABCC1), plays a critical role in the development of multidrug resistance (MDR). Ibrutinib is an inhibitor of Bruton's tyrosine kinase. Here we investigated the reversal effect of ibrutinib on MRP1-mediated MDR.
Experimental approach:
Cytotoxicity was determined by MTT assay. The expression of protein was detected by Western blot. RT-PCR and Q-PCR were performed to detect the expression of MRP1 mRNA. The intracellular accumulation and efflux of substrates for MRP1 were measured by scintillation counter and flow cytometry. HEK293/MRP1 cell xenografts in nude mice were established to study the effects of ibrutinib in vivo.
Key results:
Ibrutinib significantly enhanced the cytotoxicity of MRP1 substrates in HEK293/MRP1 and HL60/Adr cells overexpressing MRP1. Furthermore, ibrutinib increased the accumulation of substrates in these MRP1-overexpressing cells by inhibiting the drug efflux function of MRP1. However, mRNA and protein expression of MRP1 remained unaltered after treatment with ibrutinib in MRP1-overexpressing cells. In vivo, ibrutinib enhanced the efficacy of vincristine to inhibit the growth of HEK293/MRP1 tumour xenografts in nude mice. Importantly, ibrutinib also enhances the cytotoxicity of vincristine in primary cultures of leukaemia blasts, derived from patients.
Conclusions and implications:
Our results indicated that ibrutinib significantly increased the efficacy of the chemotherapeutic agents which were MRP1 substrates, in MRP1-overexpressing cells, in vitro, in vivo and ex vivo. These findings will lead to further studies on the effects of a combination of ibrutinib with chemotherapeutic agents in cancer patients overexpressing MRP1.
The COVID-19 pandemic that started in 2019 and resulted in significant morbidity and mortality continues to be a significant global health challenge, characterized by inflammation, oxidative stress, and immune system dysfunction.. Developing therapies for preventing or treating COVID-19 remains an important goal for pharmacology and drug development research. Polyphenols are effective against various viral infections and can be extracted and isolated from plants without losing their therapeutic potential. Researchers have developed methods for separating and isolating polyphenols from complex matrices. Polyphenols are effective in treating common viral infections, including COVID-19, and can also boost immunity. Polyphenolic-based antiviral medications can mitigate SARS-CoV-2 enzymes vital to virus replication and infection. Individual polyphenolic triterpenoids, flavonoids, anthraquinonoids, and tannins may also inhibit the SARS-CoV-2 protease. Polyphenol pharmacophore structures identified to date can explain their action and lead to the design of novel anti-COVID-19 compounds. Polyphenol-containing mixtures offer the advantages of a well-recognized safety profile with few known severe side effects. However, studies to date are limited, and further animal studies and randomized controlled trials are needed in future studies. The purpose of this study was to review and present the latest findings on the therapeutic impact of plant-derived polyphenols on COVID-19 infection and its complications. Exploring alternative approaches to traditional therapies could aid in developing novel drugs and remedies against coronavirus infection.
Heterocyclic compounds are well‐known for their use in the synthesis of drugs and drug intermediates. This review provides a systematic survey on the chemical synthesis of complex heterocyclic compounds containing multi‐stereocentres using 3‐arylidene‐4‐chromanones in the last two decades. Regioselective and stereoselective synthesis of complex heterocycles involving 3‐arylidene‐chromanones and 3‐arylidene‐thiochormanones are dealt under various reaction headings such as epoxidation, Michael addition, 1,3‐dipolar cycloaddition, [3+2] and [4+2] cycloaddition, Diels‐Alder reaction, sulfa‐Michael/aldol cascade reactions, asymmetric synthesis employing bifunctional catalysts, chiral phase‐transfer catalysts, chiral squaramide catalysts, etc. These synthetic highlights demonstrate the utility of 3‐arylidene‐4‐chromanones and 3‐arylidene‐thiochormanones as versatile synthons for rapid assemblage of complex condensed heterocyles and spiro ‐fused heterocycles towards the development of pharmaceutically active compounds. A brief highlight on the synthetic methods of 3‐arylidene‐4‐chromanone derivatives is also dealt. This review creates a platform for further improvement on the skeletal design of potential drug candidates.
Multitarget datasets that correlate bioactivity landscapes of small-molecules toward different related or unrelated pharmacological targets are crucial for novel drug design and discovery. ATP-binding cassette (ABC) transporters are critical membrane-bound transport proteins that impact drug and metabolite distribution in human disease as well as disease diagnosis and therapy. Molecular-structural patterns are of the highest importance for the drug discovery process as demonstrated by the novel drug discovery tool ‘computer-aided pattern analysis’ (‘C@PA’). Here, we report a multitarget dataset of 1,167 ABC transporter inhibitors analyzed for 604 molecular substructures in a statistical binary pattern distribution scheme. This binary pattern multitarget dataset (ABC_BPMDS) can be utilized for various areas. These areas include the intended design of (i) polypharmacological agents, (ii) highly potent and selective ABC transporter-targeting agents, but also (iii) agents that avoid clearance by the focused ABC transporters [ e.g ., at the blood-brain barrier (BBB)]. The information provided will not only facilitate novel drug prediction and discovery of ABC transporter-targeting agents, but also drug design in general in terms of pharmacokinetics and pharmacodynamics.
The adenosine-triphosphate-(ATP)-binding cassette (ABC) transporter ABCA7 is a genetic risk factor for Alzheimer’s disease (AD). Defective ABCA7 promotes AD development and/or progression. Unfortunately, ABCA7 belongs to the group of ‘under-studied’ ABC transporters that cannot be addressed by small-molecules. However, such small-molecules would allow for the exploration of ABCA7 as pharmacological target for the development of new AD diagnostics and therapeutics. Pan-ABC transporter modulators inherit the potential to explore under-studied ABC transporters as novel pharmacological targets by potentially binding to the proposed ‘multitarget binding site’. Using the recently reported cryo-electron microscopy (cryo-EM) structures of ABCA1 and ABCA4, a homology model of ABCA7 has been generated. A set of novel, diverse, and potent pan-ABC transporter inhibitors has been docked to this ABCA7 homology model for the discovery of the multitarget binding site. Subsequently, application of pharmacophore modelling identified the essential pharmacophore features of these compounds that may support the rational drug design of innovative diagnostics and therapeutics against AD.
Onion or Allium cepa (A. cepa) is one of the most important condiment plants grown and consumed all over the world. This plant has various therapeutic effects attributed to its constituents, such as quercetin, thiosulphinates and phenolic acids. In the present article, various pharmacological and therapeutic effects of A. cepa were reviewed. Different online databases using keywords such as onion, A. cepa, therapeutic effects, and pharmacological effects until the end of December 2019 were searched for this purpose. Onion has been suggested to be effective in treating a broad range of disorders, including asthma, inflammatory disorders, dysentery, wounds, scars, keloids and pain. In addition, different studies have demonstrated that onion possesses numerous pharmacological properties, including anti-cancer, anti-diabetic and anti-platelet properties as well as the effect on bone, cardiovascular, gastrointestinal, nervous, respiratory, and urogenital systems effects such as anti-osteoporosis, anti-hypertensive, antispasmodic, anti-diarrheal, neuro-protective, anti-asthmatic and diuretic effects. The present review provides detailed the various pharmacological properties of onion and its constituents and possible underlying mechanisms. The results of multiple studies suggested the therapeutic effect of onion on a wide range of disorders.
Onions are a widely cultivated and consumed vegetable, and contain various bioactive components, which possess various health benefits, such as antioxidant, anti-obesity, and anti-diabetic properties. As the major bioactive constituents in onions, it is essential to study phenolic compounds and the heath beneficial properties of onion and its by-products. The bioactivity of phenolic components in onions can be affected by many factors, including the genotype, different growing environments and food processing methods. Currently, most reviews have focused on an investigation of the chemical compounds or bioactivity of raw onions, but there is a paucity of studies concerning whether pre-harvest (i.e., genotype and growing environment) and post-harvest (i.e., storage) factors can impact its phenolic compounds. This review provides knowledge and guidance to agricultural production on producing high-quality onions and to the food industry on developing functional foods to reduce some chronic diseases such as diabetes. It also promotes research interest in studying bioactive compounds in fruits and vegetables considering different pre-harvest and post-harvest conditions.
Numerous studies have reported the emergence of antimicrobial resistance during the treatment of common infections. Multidrug resistance (MDR) leads to failure of antimicrobial treatment, prolonged illness, and increased morbidity and mortality. Overexpression of multidrug resistance proteins (MRPs) as drug efflux pumps are one of the main contributions of MDR, especially multidrug resistance protein 4 (MRP4/ABCC4) in the development of antimicrobial resistance. The molecular mechanism of antimicrobial resistance is still under investigation. Various intervention strategies have been developed for overcoming MDR, but the effect is limited. Suppression of MRP4 may be an attractive therapeutic approach for addressing drug resistance. However, there are few reports on the involvement of MRP4 in antimicrobial resistance and inflammatory diseases. In this review, we introduced the function and regulation of MRP4, and then summarized the roles of MRP4 in microbial infections and inflammatory diseases as well as polymorphisms in the gene encoding this transporter. Further studies should be conducted on drug therapy targeting MRP4 to improve the efficacy of antimicrobial therapy. This review can provide useful information on MRP4 for overcoming antimicrobial resistance and anti-inflammatory therapy.
Cancer causes considerable morbidity and mortality across the world. Socioeconomic, environmental, and lifestyle factors contribute to the increasing cancer prevalence, bespeaking a need for effective prevention and treatment strategies. Phytochemicals like plant polyphenols are generally considered to have anticancer, anti-inflammatory, antiviral, antimicrobial, and immunomodulatory effects, which explain their promotion for human health. The past several decades have contributed to a growing evidence base in the literature that demonstrate ability of polyphenols to modulate multiple targets of carcinogenesis linking models of cancer characteristics (i.e., hallmarks and nutraceutical-based targeting of cancer) via direct or indirect interaction or modulation of cellular and molecular targets. This evidence is particularly relevant for the lignans, an ubiquitous, important class of dietary polyphenols present in high levels in food sources such as flaxseed. Literature evidence on lignans suggests potential benefit in cancer prevention and treatment. This review summarizes the relevant chemical and pharmacokinetic properties of dietary polyphenols and specifically focuses on the biological targets of flaxseed lignans. The consolidation of the considerable body of data on the diverse targets of the lignans will aid continued research into their potential for use in combination with other cancer chemotherapies, utilizing flaxseed lignan-enriched natural products.
Several mechanisms of pharmacokinetic, metabolic, and regulatory nature have been elucidated to take part or act in concert in the phenomenon of multidrug resistance (MDR). MDR is characterized by cross‐resistance of cells against chemotherapeutic agents, which are used for treatment of e.g., cancer, bacterial infections, or human immunodeficiency virus (HIV) infections. One group of proteins that combines all three stated aspects—the metabolism and distribution of drugs as well as their own regulation—is adenosine triphosphate‐binding cassette (ABC) transporters. These efflux pumps use the energy of adenosine triphosphate hydrolysis for drug translocation from the membrane and the cytosol to the extracellular space, often with cotransport of a cosubstrate. Multidrug resistance‐associated protein 1 (MRP1, ABCC1) had been discovered as one major key player in cancer‐related MDR. The xenobiotic substrates include anthracyclines, vinca alkaloids, podophyllotoxins, as well as glutathione (GSH)‐adducts of certain cytostatics. Contrary to other transport proteins involved in cancer‐related MDR the activity of MRP1 is related to the GSH content of cells. A modern strategy to overcome MRP1‐associated MDR is besides its inhibition the activation of GSH efflux, enforcing cell death due to cellular stress. In addition, it has recently been found that MRP1 contributes to the β‐amyloid protein clearance in Alzheimer's disease (AD). Collectively, transport activation of MRP1 is of therapeutic value, and furthermore helps to elucidate the transport protein function and the mechanisms behind it. This review is meant to summarize the known concepts of MRP1 activation, which might contribute to a further understanding of MRP1 in particular and ABC transporters in general.
Quercetin is a biologically active flavonoid that has been used as a popular health supplement. It is reported that quercetin may cause flavonoid-drug interaction mediated by P-glycoprotein, the most predominant efflux transporter. In this study, we comprehensively evaluated the potential of the pharmacokinetic interaction of quercetin mediated by multidrug resistance-associated protein 2 (MRP2), another major efflux transporter. MRP2-transfected MDCKII cells and LS174T cells were used to evaluate the potential inhibition and induction of MRP2 by quercetin in vitro. To evaluate the induction effect of quercetin on Mrp2 in vivo, Mrp2 mRNA expression in rat liver, kidney, and small intestinal tissues was determined after the oral administration of quercetin (50, 100, or 250 mg/kg) for seven days. Mrp2-mediated interaction potential was also evaluated by the pharmacokinetic study of phenolsulfonphthalein in rats after single or multiple doses of quercetin. Additionally, the effect of quercetin on absorption of docetaxel, a P-glycoprotein and CYP3A4 substrate, was also evaluated. Quercetin inhibited the function of MRP2 at 10 µM and induced the mRNA expression of MRP2 at 50 µM in vitro. Additionally, at 100 mg/kg, quercetin markedly increased Mrp2 expression in the small intestine of rats. However, there was no significant change in phenolsulfonphthalein pharmacokinetics due to single- (50, 100, or 250 mg/kg) or multiple-dose (50, 100, or 250 mg/kg for seven days) quercetin co-administration. By contrast, a significant interaction caused by quercetin (100 mg/kg) was observed in the absorption of docetaxel. The results suggested that although quercetin modulates the function and expression of MRP2 in vitro, it may have a low potential of Mrp2-mediated interaction and present negligible safety concerns related to the interaction.
Foxtail millet is the second-most widely planted species of millet and the most important cereal food in China. Our previous study showed that bound polyphenol of inner shell (BPIS) from foxtail millet bran displayed effective anti-tumor activities in vitro and in vivo. The present research further implied that BPIS has the ability to reverse the multi-drug resistance of colorectal cancer in human HCT-8/Fu cells, the IC50 values of 5-fluorouracil (5-Fu), oxaliplatin (L-OHP) and vincristine (VCR) were decreased form 6593 ± 53.8 , 799 ± 48.9 , 247 ± 10.3 μM to 5350 ± 22.3 (3261 ± 56.9) , 416 ± 16.6 (252 ± 15.6) and 144 ± 8.30 (83.8 ± 5.60) μM when HCT-8/Fu cells pretreated with 0.5 (1.0) mg/ml BPIS for 12 h. The 12 phenolic acid compounds of BPIS were identified by UPLC-Triple-TOF/MS method. Especially, the fraction of molecular weight (MW) < 200 of BPIS reversed the multi-drug resistance in HCT-8/Fu cells, and ferulic acid and p-coumaric acid were the main active components, the IC50 values were 1.23 ± 0.195 and 2.68 ± 0.163 mg/ml , respectively. The present data implied that BPIS significantly enhanced the sensitivity of chemotherapeutic drugs through inhibiting cell proliferation, promoting cell apoptosis and increasing the accumulation of rhodamine-123 (Rh-123) in HCT-8/Fu cells. RT-PCR and western blot data indicated that BPIS also decreased the expression levels of multi-drug resistance protein 1 (MRP1), P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). Collectively, these results show that BPIS has potential ability to be used as a new drug-resistance reversal agent in colorectal cancer.
Nature's providences are rather the choicest remedies for human health and welfare. One such is quercetin, which is nature's nominee for cancer cure and recently demonstrated against influenza attack. Quercetin is highly recognized for its anticancer applications. This review emphasizes on yet another gift that this compound has to offer for mankind, which is none other than combating the deadly evasive influenza virus. The chemistry of this natural bioflavonoid and its derivatives and its modus operandi against influenza virus is consolidated into this review. The advancements and achievements made in the anti-influenza clinical history are also documented. Further, the challenges facing the progress of this compound to emerge as a predominant anti-influenza drug are discussed, and the future perspective for breaking its limitations through integration with nanoplatforms is envisioned.
Arsenic trioxide (ATO) is highly efficient in treating acute promyelocytic leukemia. Other malignancies, however, are often less sensitive. Searching for compounds sensitizing arsenic resistant tumours for ATO the plant polyphenols, chrysin and silibinin, and the ATP binding cassette (ABC) transporter inhibitor MK-571, respectively, were investigated in human glioblastoma A-172 cells. The sensitivity of A-172 cells to ATO was characterized by a median cytotoxic concentration of 6 μM ATO. Subcytotoxic concentrations of chrysin, silibinin and MK-571, respectively, remarkably increased the sensitivity of the cells to ATO by factors of 4–6. Isobolographic analysis revealed synergistic interaction of the polyphenols and MK-571, respectively, with ATO. Sensitization by chrysin was associated with depletion of cellular glutathione and increased accumulation of arsenic. In contrast, silibinin and also MK-571 increased the accumulation of arsenic more strongly but without affecting the glutathione level. The increase of arsenic accumulation could be attributed to a decreased rate of arsenic export and, additionally, in the case of silibinin and MK-571, to an increasing amount of irreversibly accumulated arsenic. Direct interaction with ABC transporters stimulating export of glutathione and inhibiting export of arsenic, respectively, are discussed as likely mechanisms of the sensitizing activity of chrysin and silibinin.
Background:
Ageing can be simply defined as the process of becoming older, which is genetically determined but also environmentally modulated. With the continuous increase of life expectancy, quality of life during ageing has become one of the biggest challenges of developed countries. The quest for a healthy ageing has led to the extensive study of plant polyphenols with the aim to prevent age-associated deterioration and diseases, including neurodegenerative diseases. The world of polyphenols has fascinated researchers over the past decades, and in vitro, cell-based, animal and human studies have attempted to unravel the mechanisms behind dietary polyphenols neuroprotection.
Methods:
In this review, we compiled some of the extensive and ever-growing research in the field, highlighting some of the most recent trends in the area.
Results:
The main findings regarding (poly)phenols neuroprotective potential performed using in vitro, cellular and animal studies, as well as human trials are covered in this review. Concepts like bioavailability, polyphenols biotransformation, transport of dietary polyphenols across barriers, including the blood-brain barrier, are here explored.
Conclusion:
The diversity and holistic properties of (poly)phenol present them as an attractive alternative for the treatment of multifactorial diseases, where a multitude of cellular pathways are disrupted. The underlying mechanisms of (poly)phenols for nutrition or therapeutic applications must be further consolidated, however there is strong evidence of their beneficial impact on brain function during ageing. Nevertheless, only the tip of the iceberg of nutritional and pharmacological potential of dietary polyphenols is hitherto understood and further research needs to be done to fill the gaps in pursuing a healthy ageing.
The high incidence of breast cancer in developed and developing countries, and its correlation to cancer-related deaths, has prompted concerned scientists to discover novel alternatives to deal with this challenge. In this review, we will provide a brief overview of polyphenol structures and classifications, as well as on the carcinogenic process. The biology of breast cancer cells will also be discussed. The molecular mechanisms involved in the anti-cancer activities of numerous polyphenols, against a wide range of breast cancer cells, in vitro and in vivo, will be explained in detail. The interplay between autophagy and apoptosis in the anti-cancer activity of polyphenols will also be highlighted. In addition, the potential of polyphenols to target cancer stem cells (CSCs) via various mechanisms will be explained. Recently, the use of natural products as chemotherapeutics and chemopreventive drugs to overcome the side effects and resistance that arise from using chemical-based agents has garnered the attention of the scientific community. Polyphenol research is considered a promising field in the treatment and prevention of breast cancer.
Membrane transporter proteins (the ABC transporters and SLC transporters) play pivotal roles in drug absorption and disposition, and thus determine their efficacy and safety. Accumulating evidence suggests that the expression and activity of these transporters may be modulated by various phytochemicals (PCs) found in diets rich in plants and herbs. PC absorption and disposition are also subject to the function of membrane transporter and drug metabolizing enzymes. PC–drug interactions may involve multiple major drug transporters (and metabolizing enzymes) in the body, leading to alterations in the pharmacokinetics of substrate drugs, and thus their efficacy and toxicity. This review summarizes the reported in vitro and in vivo interactions between common dietary PCs and the major drug transporters. The oral absorption, distribution into pharmacological sanctuaries and excretion of substrate drugs and PCs are considered, along with their possible interactions with the ABC and SLC transporters which influence these processes.
Drug resistance develops in nearly all patients with colon cancer, leading to a decrease in the therapeutic efficacies of anticancer agents. This review provides an up-to-date summary on over-expression of ATP-binding cassette (ABC) transporters and evasion of apoptosis, two representatives of transport-based and non-transport-based mechanisms of drug resistance, as well as their therapeutic strategies. Different ABC transporters were found to be up-regulated in colon cancer, which can facilitate the efflux of anticancer drugs out of cancer cells and decrease their therapeutic effects. Inhibition of ABC transporters by suppressing their protein expressions or co-administration of modulators has been proven as an effective approach to sensitize drug-resistant cancer cells to anticancer drugs in vitro. On the other hand, evasion of apoptosis observed in drug-resistant cancers also results in drug resistance to anticancer agents, especially to apoptosis inducers. Restoration of apoptotic signals by BH3 mimetics or epidermal growth factor receptor inhibitors and inhibition of cancer cell growth by alternative cell death pathways, such as autophagy, are effective means to treat such resistant cancer types. Given that the drug resistance mechanisms are different among colon cancer patients and may change even in a single patient at different stages, personalized and specific combination therapy is proposed to be more effective and safer for the reversal of drug resistance in clinics.
Allium chinense is a medicinal plant and nutritional food commonly used in Eastern Asia. In this study, we investigated the in vitro antioxidant activity (scavenging of α,α-diphenyl-β-picrylhydrazyl free radical, total phenol content, reducing power, and total antioxidant activity) and constituents of various extracts from A. chinense. Moreover, we also studied the in vivo hypolipidemic effects of extracts on high-fat-diet Wistar rats. Ethanol extracts from A. chinense showed notable antioxidant activity, and its high-dose essential-oil extract both significantly reduced serum and hepatic total cholesterol, triglyceride, and low-density lipoprotein levels and increased serum high-density lipoprotein levels in high-fat-diet Wistar rats compared with those observed following treatment with the control drug probucol. Additionally, visceral fat in high-fat-diet Wistar rats was reduced. Furthermore, groups with high doses of essential-oil and residue extracts showed protective effects associated with histopathological liver alteration. These results suggested that A. chinense is a valuable plant worthy of further investigation as a potential dietary supplement or botanical drug.
Accumulated evidences have focused on the use of natural polyphenolic compounds as nutraceuticals, since they showed a wide range of bioactivities and exhibited protection against variety of age related disorders. Polyphenols have variable potencies to interact, and hence alter the activities of various transporter proteins, many of them classified as ATP-Binding Cassette transporters, like multidrug resistance protein (MDRP), and p-glycoprotein (P-gp). Some of the efflux transporters are generally linked with anticancer and antiviral drug resistance; in this context, polyphenols may be beneficial in modulating drug resistance by increasing the efficacy of anticancer and antiviral drugs. Additionally, these effects were implicated to explain the influence of dietary polyphenols on drug efficacy as result of food-drug interactions. However, limited data are available about the influence of these components on uptake transporters. Therefore, the objective of this article is to review the potential efficacies of polyphenols in modulating the functional integrity of uptake transporter proteins, including those terminated the effect of neurotransmitters, and their possible influence in neuropharmacology.
Objective:
To evaluate the in vitro activities of the ethyl acetate (EA) fraction of Houttuynia cordata (H. cordata) Thunb. (Saururaceae) and three of its constituent flavonoids (quercetin, quercitrin and rutin) against murine coronavirus and dengue virus (DENV).
Methods:
The antiviral activities of various concentrations of the EA fraction of H. cordata and flavonoids were assessed using virus neutralization tests against mouse hepatitis virus (MHV) and DENV type 2 (DENV-2). Cinanserin hydrochloride was also tested against MHV. The EA fraction of H. cordata was tested for acute oral toxicity in C57BL/6 mice.
Results:
The EA fraction of H. cordata inhibited viral infectivity up to 6 d. Cinanserin hydrochloride was able to inhibit MHV for only 2 d. The 50% inhibitory concentrations (IC50) of the EA fraction of H. cordata added before the viral adsorption stage were 0.98 μg/mL for MHV and 7.50 μg/mL for DENV-2 with absence of cytotoxicity. The mice fed with the EA fraction up to 2000 mg/kg did not induce any signs of acute toxicity, with normal histological features of major organs. Certain flavonoids exhibited comparatively weaker antiviral activity, notably quercetin which could inhibit both MHV and DENV-2. This was followed by quercitrin which could inhibit DENV-2 but not MHV, whereas rutin did not exert any inhibitory effect on either virus. When quercetin was combined with quercitrin, enhancement of anti-DENV-2 activity and reduced cytotoxicity were observed. However, the synergistic efficacy of the flavonoid combination was still less than that of the EA fraction.
Conclusions:
The compounds in H. cordata contribute to the superior antiviral efficacy of the EA fraction which lacked cytotoxicity in vitro and acute toxicity in vivo. H. cordata has much potential for the development of antiviral agents against coronavirus and dengue infections.
32P-postlabeiling analysis indicated the time-dependent formation of five char-acteristic spots of adducts with the endogenous DNA in broccoli homogenates. One of these adduct spots was also consistently detected in tissues of rats and mice fed raw or steamed broccoli. Its structure was identified by mass spectrometry and direct chemical synthesis. It was formed by a specific glucosinolate (GLS-A); administration of purified GLS-A or a degradation product formed from GLS-A by myrosinase (DEG-A) to mice led to the formation of the same DNA adduct as observed after feeding broccoli. However, when purified GLS-A was used, adduct formation was largely restricted to the large bowel, whereas adducts were detected in many additional tissues after feeding broccoli or administering DEG-A. This difference may result from the enzymes mediating the first step of bioactivation: glycosidases from intestinal bacteria with puri-fied GLS-A, plant myrosinase in the case of broccoli; this activation step is not required with the DEG-A. Unlike other GLS, GLS-A was highly mutagenic to Salmonella typhimurium strains in the presence of myrosinase. It remains to be studied whether broccoli-induced mutagenic DNA adducts can induce tumour-igenesis.
This chapter focuses on the interactions of diet/nutrients with drug transporters arising from in vitro and in vivo studies, with specific emphasis on several major efflux transporters and uptake transporters. Most in vivo interaction studies reported to date are focused primarily on the interactions between dietary supplements and P-glycoprotein (P-gp). Given the facts that multidrug resistance associated protein (MRP) and breast cancer resistance protein (BCRP), organic anion-transporting polypeptide (OATP), and organic anion transporter (OAT) are also essential in drug disposition, it is important to appreciate the potential PK interactions of diet/nutrients with these transporters. The concentrations of many flavonoids required to produce significant modulation on activities of these transporters appear to be, in general, within the micro-molar range, which is achievable in the intestine after intake of food and, especially, after ingestion of dietary supplements.
Resistance to chemotherapeautic drugs is one of the main obstacles to effective cancer treatment. Multidrug resistance (MDR) is defined as resistance to structurally and/or functionally unrelated drugs, and has been extensively investigated for the last three decades. There are two types of MDR: intrinsic and acquired. Tumor microenvironment selection pressure leads to the development of intrinsic MDR, while acquired resistance is a consequence of the administered chemotherapy. A central issue in chemotherapy failure is the existence of heterogeneous populations of cancer cells within one patient and patient-to-patient variability within each type of cancer. Numerous genes and pathways contribute to the development of MDR in cancer. Point mutations, gene amplification or other genetic or epigenetic changes all affect biological functions and may lead to the occurrence of MDR phenotype. Similar to the characteristics of cancerogenesis, the main features of MDR include abnormal tumor vasculature, regions of hypoxia, aerobic glycolysis, and a lower susceptibility to apoptosis. In order to achieve a lethal effect on cancer cells, drugs need to reach their intracellular target molecules. The overexpression of the efflux transporter P-glycoprotein (P-gp) in MDR cancer cells leads to decreased uptake of the drug and intracellular drug accumulation, minimising drug-target interactions. New agents being or inspired by natural products that sucessfully target these mechanisms are the main subject of this review. Two key approaches in combating MDR in cancer are discussed (i) finding agents that preserve citotoxicity toward MDR cancer cells; (ii) developing compounds that restore the cytotoxic activity of classic anticancer drugs.
Multidrug-resistant protein 4 (MRP4), a member of the C subfamily of ATP-binding cassette (ABC) transporters, is distributed in a variety of tissues and a number of cancers. As a drug transporter, MRP4 is responsible for the pharmacokinetics and pharmacodynamics of numerous drugs, especially antiviral drugs, antitumor drugs and diuretics. In this regard, the functional role of MRP4 is affected by a number of factors, such as genetic mutations, tissue-specific transcriptional regulations, post-transcriptional regulations including miRNAs and membrane internalization, and substrate competition. Unlike other C family members, MRP4 is in a pivotal position to transport cellular signaling molecules, through which it is tightly connected to the living activity and physiological processes of cells and bodies. In the context of several cancers in which MRP4 is overexpressed, MRP4 inhibition shows striking effects against cancer progression and drug resistance. In this review, we describe the role of MRP4 more specifically in both healthy conditions and disease states, with an emphasis on its potential as a drug target.
The American Society for Pharmacology and Experimental Therapeutics.
The gene family of ATP-binding cassette (ABC) transporters is widely distributed over the evolutionary tree of life from bacteria to man. In cancer, several ABC transporters are involved in the transport of anticancer drugs. This leads to the development of resistance to a broad spectrum of anticancer agents, termed multidrug resistance. An attractive strategy to overcome multidrug resistance is to block the transport function of ABC transporters leading to lethal intracellular concentrations of anticancer drugs. Efforts to identify transport inhibitors lead to a huge amount of chemical substances, none of which successfully passed clinical trials in cancer patients because of high toxicity. The search of natural products from traditional Chinese medicine may be more promising because natural products frequently are less toxic than chemically synthesized xenobiotics. Here, we give an overview of ABC transporters involved in multidrug resistance of cancer as well as Chinese herbs and phytochemicals showing inhibitory activity towards ABC transporters.
Over 80% of ovarian cancer patients develop chemoresistance which results in a lethal course of the disease. A well-established cause of chemoresistance involves the family of ATP-binding cassette transporters, or ABC transporters that transport a wide range of substrates including metabolic products, nutrients, lipids, and drugs across extra- and intra-cellular membranes. Expressions of various ABC transporters, shown to reduce the intracellular accumulation of chemotherapy drugs, are increased following chemotherapy and impact on ovarian cancer survival. Although clinical trials to date using ABC transporter inhibitors have been disappointing, ABC transporter inhibition remains an attractive potential adjuvant to chemotherapy. A greater understanding of their physiological functions and role in ovarian cancer chemoresistance will be important for the development of more effective targeted therapies. This article will review the role of the ABC transporter family in ovarian cancer progression and chemoresistance as well as the clinical attempts used to date to reverse chemoresistance.
Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.
Multidrug resistance is one of the most common causes of relapse in cancer chemotherapy. Inhibition of ABC transporters to reverse MDR is a prominent approach to enhance the efficacy of cancer chemotherapy. We investigated the effect of resveratrol (RSV) on the membrane transport function and the expression of proteins involved in the multidrug resistance in NCI-H460 cells. The molecular interactions of RSV with P-gp were analyzed by Schrodinger software. The membrane transport function and cell cycle distribution were measured using flow cytometry. The mRNA expression level of MDR1, LRP, MRP2, ABCC1, ABCC2 and ABCC3 genes were detected by qRT-PCR and BCRP expression was detected by western blot analysis. In silico docking studies revealed that RSV possesses greater binding affinity with TMD region of P-gp. In this study, RSV pretreatment significantly enhanced Paclitaxel (PTX) antiproliferative effect in NCI-H460 cells. The rhodamine 123 drug efflux studies revealed that there was a significant transport function inhibition by RSV treatment and moderate transport function inhibition by PTX. Further, RSV treatment significantly decreased the mRNA expression levels of various ABC transporters genes. Furthermore, expression of BCRP was found to be down-regulated during RSV treatment. It was also found that this enhanced anticancer efficacy of RSV was associated with PTX-induced cell arrest in the G2/M phase of cell cycle. Interestingly, we observed significantly enhanced antiproliferative effect, transport function inhibition and downregulation of ABC transporters in RSV-PTX combination group. This might be due to additive or synergistic effect of RSV with PTX in NCI-H460 cells. Thus, the present findings illustrate the modulatory role of RSV on PTX sensitization in relatively resistant NCI-H460 cells.
Multidrug resistance (MDR) is a condition that makes cells simultaneously unresponsive to different drugs, unrelated to their chemical structure and mechanism of action. MDR caused by the presence and overexpression of ABC transporters makes obstacles in cancer treatment and lower the effectiveness of chemotherapy. Natural products are investigated by many researchers as MDR modulators for their low toxicity and potent, selective behavior. When coadministered, MDR modulators compete with cytotoxic agents for binding to the active site of the membrane transporters and reduce drug efflux. Natural product-based drugs are important in struggling against drug resistance during cancer therapy. This review is focused on the potential mechanisms against drug resistance, the development of inhibitors for ABC drug transporters, natural product modulators, and nanoparticle drug delivery.
Neuroblastoma is the most common cancer of infancy and accounts for 15% of all pediatric oncology deaths. Survival rates of high-risk neuroblastoma remain less than 50%, with amplification of the MYCN oncogene the most important aberration associated with poor outcome. Direct transcriptional targets of MYCN include a number of ATP-binding cassette (ABC) transporters, of which ABCC1 (MRP1), ABCC3 (MRP3), and ABCC4 (MRP4) are the best characterized. These three transporter genes have been shown to be strongly prognostic of neuroblastoma outcome in primary untreated neuroblastoma. In addition to their ability to efflux a number of chemotherapeutic drugs, evidence suggests that these transporters also contribute to neuroblastoma outcome independent of any role in cytotoxic drug efflux. Endogenous substrates of ABCC1 and ABCC4 that may be potential candidates affecting neuroblastoma biology include molecules such as prostaglandins and leukotrienes. These bioactive lipid mediators have the ability to influence biological processes contributing to cancer initiation and progression, such as angiogenesis, cell signaling, inflammation, proliferation, and migration and invasion. ABCC1 and ABCC4 are thus potential targets for therapeutic suppression in high-risk neuroblastoma, and recently developed small-molecule inhibitors may be an effective strategy in treating aggressive forms of this cancer, as well as other cancers that express high levels of these transporters.
© 2015 Elsevier Inc. All rights reserved.
Conventional cancer chemotherapy is seriously limited by tumor cells exhibiting multidrug resistance (MDR), which is caused by changes in the levels or activity of membrane transporters that mediate energy-dependent drug efflux and of proteins that affect drug metabolism and/or drug action. Cancer scientists and oncologists have worked together for some time to understand anticancer drug resistance and develop pharmacological strategies to overcome such resistance. Much focus has been on the reversal of the MDR phenotype by inhibition of ATP-binding cassette (ABC) drug transporters. ABC transporters are a family of transporter proteins that mediate drug resistance and low drug bioavailability by pumping various drugs out of cells at the expense of ATP hydrolysis. Many inhibitors of MDR transporters have been identified, and though some are currently undergoing clinical trials, none are in clinical use. Herein, we briefly review the status of MDR in human cancer, explore the pathways of MDR in chemotherapy, and outline recent advances in the design and development of MDR modulators.
The blood-brain barrier (BBB) is a dynamic and complex interface between the blood and the central nervous system regulating brain homeostasis. Major functions of the BBB include the transport of nutrients and protection of the brain from toxic compounds. This review summarizes the most important transport pathways contributing to the nutrition of the brain. Carrier-mediated transport selectively delivers small molecules like sugars, amino acids, vitamins, and trace elements. Large biomolecules, lipoproteins, peptide and protein hormones cross the BBB by receptor-mediated transport. Active efflux transporters participate in the brain efflux of endogenous metabolites as well as toxins, xenobiotics and drugs. Dysfunction in the transport of nutrients at the BBB is described in several neurological disorders and diseases. The BBB penetration of neuroprotective nutrients, especially plant polyphenols and alkaloids, their potential protective effect on brain endothelium and the interaction of nutraceuticals with active efflux transporters at the BBB are discussed. In vitro BBB models to examine nutrient transport are also presented.
Copyright © 2014 IMSS. Published by Elsevier Inc. All rights reserved.
Multidrug resistance protein 4 (MRP4) effluxes a wide variety of drugs and endogenous signaling molecules from cells and has been proposed as an attractive therapeutic target in several solid tumors, including neuroblastoma and colorectal cancer. MRP4 also regulates the pharmacokinetics of its drug substrates and its absence can increase their tissue penetration. We observed that MRP4 can efflux the bioluminescence substrate d-luciferin, and exploited this phenomenon to develop a robust, high throughput, live cell-based bioluminescent screen to identify new MRP4 inhibitors. We applied this screen to a combined library of 3600 compounds, all of which were either FDA-approved drugs or bioactive compounds with defined mechanisms of action. From the primary screen, 36 compounds effectively inhibited MRP4 (>4-fold increase in bioluminescence), with inhibitors of receptor tyrosine kinases and phosphodiesterases highly over-represented. Selected compounds were tested for their ability to sensitize MRP4-overexpressing cell lines to the MRP4 substrate drugs 6-mercaptopurine and SN-38, with sensitization up to 6.5-fold with the ryanodine receptor antagonist dantrolene. These newly identified MRP4 inhibitors are readily available and are either established drugs or well-characterized bioactive compounds. As such, they should be immediately useful as investigative tools, and suitable for testing both in vitro and in vivo.
Copyright © 2014 Elsevier Inc. All rights reserved.
Chemotherapy resistance represents a major problem for the treatment of patients with breast cancer and greatly restricts the use of first-line chemotherapeutics paclitaxel. The purpose of this study was to investigate the role of transgelin 2 in human breast cancer paclitaxel resistance cell line (MCF-7/PTX) and the reversal mechanism of salvianolic acid A (SAA), a phenolic active compound extracted from Salvia miltiorrhiza. Western blotting and real-time quantitative polymerase chain reaction (qRT-PCR) indicated that transgelin 2 may mediate paclitaxel resistance by activating the phosphatidylinositol 3-kinase (PI3 K)/Akt signaling pathway to suppress MCF-7/PTX cells apoptosis. The reversal ability of SAA was confirmed by MTT assay and flow cytometry, with a superior 9.1-fold reversal index and enhancement of the apoptotic cytotoxicity induced by paclitaxel. In addition, SAA effectively prevented transgelin 2 and adenosine-triphosphate binding cassette transporter (ABC transporter) including P-glycoprotein (P-gp), multidrug resistance associated protein 1 (MRP1), and breast cancer resistance protein (BCRP) up-regulation and exhibited inhibitory effect on PI3 K/Akt signaling pathway in MCF-7/PTX cells. Taken together, SAA can reverse paclitaxel resistance through suppressing transgelin 2 expression by mechanisms involving attenuation of PI3 K/Akt pathway activation and ABC transporter up-regulation. These results not only provide insight into the potential application of SAA in reversing paclitaxel resistance, thus facilitating the sensitivity of breast cancer chemotherapy, but also highlight a potential role of transgelin 2 in the development of paclitaxel resistance in breast cancer.
Copyright © 2014 Elsevier GmbH. All rights reserved.
The main dietary sources of polyphenols are reviewed, and the daily intake is calculated for a given diet containing some common fruits, vegetables and beverages. Phenolic acids account for about one third of the total intake and flavonoids account for the remaining two thirds. The most abundant flavonoids in the diet are flavanols (catechins plus proanthocyanidins), anthocyanins and their oxidation products. The main polyphenol dietary sources are fruit and beverages (fruit juice, wine, tea, coffee, chocolate and beer) and, to a lesser extent vegetables, dry legumes and cereals. The total intake is ∼1 g/d. Large uncertainties remain due to the lack of comprehensive data on the content of some of the main polyphenol classes in food. Bioavailability studies in humans are discussed. The maximum concentration in plasma rarely exceeds 1 μM after the consumption of 10–100 mg of a single phenolic compound. However, the total plasma phenol concentration is probably higher due to the presence of metabolites formed in the body's tissues or by the colonic microflora. These metabolites are still largely unknown and not accounted for. Both chemical and biochemical factors that affect the absorption and metabolism of polyphenols are reviewed, with particular emphasis on flavonoid glycosides. A better understanding of these factors is essential to explain the large variations in bioavailability observed among polyphenols and among individuals.
We have previously identified and characterized a novel member of the ATP-binding cassette superfamily of transport proteins, multidrug resistance protein (MRP), and subsequently demonstrated that its overexpression is sufficient to confer multidrug resistance on previously sensitive cells (Cole et al., Science (Washington DC), 258: 1650-1654, 1992; Grant et al., Cancer Res. 54: 357-361, 1994). In the present study, we have transfected two different eukaryotic expression vectors containing MRP complementary DNA into HeLa cells to study the pharmacological phenotype produced exclusively by overexpression of human MRP. The drug resistance patterns of the two MRP-transfected cell populations were similar. They were characterized by a moderate (5- to 15-fold) level of resistance to doxorubicin, daunorubicin, epirubicin, vincristine, and etoposide, and a low (< or = 3-fold) level of resistance to taxol, vinblastine, and colchicine. The transfectants were not resistant to 9-alkyl anthracyclines, mitoxantrone, or cisplatin. The MRP-transfected cells were also resistant to some heavy metal anions including arsenite, arsenate, and trivalent and pentavalent antimonials but were not resistant to cadmium chloride. Accumulation of radiolabeled vincristine was reduced by 45% in the MRP-transfected cells and could be restored to the levels found in sensitive cells by depletion of ATP. Rates of vincristine efflux did not differ greatly in the sensitive and resistant cells. The cytotoxic effects of vincristine and doxorubicin could be enhanced in a dose-dependent fashion by coadministration of verapamil. Cyclosporin A also increased vincristine toxicity but had less effect on doxorubicin toxicity. The degree of chemosensitization by verapamil and cyclosporin A was similar in MRP-transfected cells and in cells transfected with the vector alone, suggesting that sensitization involved mechanisms independent of MRP expression. Verapamil and cyclosporin A caused a modest increase in vincristine accumulation in the resistant cells but did not restore levels to those of the sensitive cells. Taken together, these data indicate that drug-resistant cell lines generated by transfection with MRP complementary DNA display some but not all of the characteristics of MRP-overexpressing cell lines produced by drug selection in vitro. They further demonstrate that the multidrug resistance phenotype conferred by MRP is similar but not identical to that conferred by P-glycoprotein and includes resistance to arsenical and antimonial oxyanions.
Amplification of the gene encoding multidrug resistance-associated protein (MRP) and overexpression of its cognate mRNA have been detected in multidrug-resistant cell lines derived from several different tumor types. To establish whether or not the increase in MRP is responsible for drug resistance in these cell lines, we have transfected HeLa cells with MRP expression vectors. The transfectants display an increase in resistance to doxorubicin that is proportional to the levels of a M(r) 190,000, integral membrane protein recognized by anti-MRP antibodies. The transfectants are also resistant to vincristine and VP-16 but not to cisplatin. The results demonstrate that MRP overexpression confers a multidrug resistance phenotype similar to that formerly associated exclusively with elevated levels of P-glycoprotein.
Considerable uncertainty surrounds the stoichiometry of coupling of ATP hydrolysis to drug pumping by P-glycoprotein, the multidrug transporter. To estimate relative turnovers for pumping of the drug vinblastine and ATP hydrolysis, we began by measuring the number of P-glycoprotein molecules on the surface of murine NIH3T3 cells expressing the human MDR1 gene. Fluorescence of cells treated with monoclonal antibody UIC2 was determined as a function of (i) amount of antibody at a fixed number of cells and (ii) increasing cell number at constant antibody. The two together gives 1.95 x 10(6) P-glycoprotein molecules/cell. Initial uptake rates of vinblastine +/- verapamil measure the ability of P-glycoprotein to extract vinblastine from the plasma membrane before it enters the cell. As a function of [vinblastine] at 37 degrees C, they give the maximum rate of this component of outward pumping as 2.1 x 10(6) molecules s-1 cell-1 or a turnover number of 1.1 s-1. Initial rates of one-way efflux as a function of [vinblastine] at 25 degrees C +/- glucose give the maximum rate of this component of pumping as 0.59 x 10(6) molecules s-1 cell-1. The ratio of ATPase activity of P-glycoprotein at 37 and 25 degrees C is 4.6. Appropriating this ratio for pumping, maximum one-way efflux at 37 degrees C is 4.6 x 0.59 = 2.7 x 10(6) molecules s-1 cell-1, a turnover number of 1.4 s-1. The vinblastine-stimulated ATPase activity of P-glycoprotein has a turnover number of 3.5 s-1 at 37 degrees C, giving 2.8 molecules of ATP hydrolyzed for every vinblastine molecule transported in a particular direction. These calculations involve several approximations, but turnover numbers for pumping of vinblastine and for vinblastine-stimulated ATP hydrolysis are comparable. Thus, ATP hydrolysis is probably directly linked to drug transport by P-glycoprotein.
Considerable evidence has accumulated indicating that the multidrug transporter or P-glycoprotein plays a role in the development of simultaneous resistance to multiple cytotoxic drugs in cancer cells. In recent years, various approaches such as mutational analyses and biochemical and pharmacological characterization have yielded significant information about the relationship of structure and function of P-glycoprotein. However, there is still considerable controversy about the mechanism of action of this efflux pump and its function in normal cells. This review summarizes current research on the structure-function analysis of P-glycoprotein, its mechanism of action, and facts and speculations about its normal physiological role.
Several members of the ABC family of proteins have been implicated in multidrug resistance associated with cancer therapies. A novel member of this gene family, designated pABC11, has been identified using degenerate polymerase chain reaction. The full-length cDNA spans 5881 base pairs and encodes an open reading frame of 1437 amino acids predicted to contain two sets of transmembrane domains and two nucleotide binding domains characteristic of ABC proteins. The nucleotide sequence described herein extends that of three recently reported sequences, MRP5 (Kool, M., de Haas, M., Scheffer, G., Scheper, R., van Eijk, M., Juijn, J., Baas, F., and Borst, P. (1997) Cancer Res. 57, 3537-3547), SMRP (Suzuki, T., Nishio, K., Sasaki, H., Kurokawa, H., Saito-Ohara, F., Ikeuchi, T., Tanabe, S., Terada, M., and Saijo, N. (1997) Biochem. Biophys. Res. Commun. 238, 790-794), and MOAT-C (Belinsky, M., Bain, L., Balsara, B., Testa, J., and Kruh, G. (1998) J. Natl. Cancer Inst. 90, 1735-1741), in the 5' direction. Northern blot analysis detected five transcripts that were differentially expressed in several tissue types, and the gene encoding pABC11 was mapped to chromosome 3. Confocal imaging of HEK293 cells expressing a green fluorescent protein-pABC11 construct confirmed plasma membrane localization of the fusion protein. Overexpression of pABC11 resulted in reduced labeling with the fluorochromes 5-chloromethylfluorescein diacetate, fluorescein diacetate, and 2',7'-bis-(2-carboxyethyl)-5 (and-6)-carboxyfluorescein acetoxymethyl ester but not with calcein or rhodamine derivatives, consistent with pABC11 being an anion transporter. Fluorochrome export was ATP-dependent but glutathione-independent. We also show that this export pump does not confer resistance to various classes of cytotoxic drugs but does provide small but significant resistance to CdCl(2) and potassium antimonyl tartrate.
Dideoxynucleosides, which are potent inhibitors of HIV reverse transcriptase and other viral DNA polymerases, are a common component of highly active anti-retroviral therapy (HAART) (ref. 1). Six reverse transcriptase inhibitors have been approved for human use: azidothymidine; 2'3'-dideoxycytidine; 2'3'-dideoxyinosine; 2',3'-didehydro-3'deoxythymidine; 2',3'-dideoxy-3'-thiacytidine; and 4-[2-amino-6-(cyclopropylamino)
-9H-purin-9-yl]-2-cyclopentene-1-methanol. Although drug-resistant HIV strains resulting from genetic mutation have emerged in patients treated with HAART (ref. 1), some patients show signs of drug resistance in the absence of drug-resistant viruses2, ³. In our study of alternative or additional mechanisms of resistance operating during antiviral therapy, overexpression and amplification of the MRP4 gene correlated with ATP-dependent efflux of PMEA (9-(2-phosphonylmethoxyethyl)adenine) and azidothymidine monophosphate from cells and, thus, with resistance to these drugs. Overexpression of MRP4 mRNA and MRP4 protein severely impaired the antiviral efficacy of PMEA, azidothymidine and other nucleoside analogs. Increased resistance to PMEA and amplification of the MRP4 gene correlated with enhanced drug efflux; transfer of chromosome 13 containing the amplified MRP4 gene conferred resistance to PMEA. MRP4 is the first transporter, to our knowledge, directly linked to the efflux of nucleoside monophosphate analogs from mammalian cells.
Two prominent members of the ATP-binding cassette superfamily of transmembrane proteins, multidrug resistance 1 (MDR1) P-glycoprotein and multidrug resistance protein 1 (MRP1), can mediate the cellular extrusion of xenobiotics and (anticancer) drugs from normal and tumor cells. The MRP subfamily consists of at least six members, and here we report the functional characterization of human MRP5. We found resistance against the thiopurine anticancer drugs, 6-mercaptopurine (6-MP) and thioguanine, and the anti-HIV drug 9-(2-phosphonylmethoxyethyl)adenine (PMEA) in MRP5-transfected cells. This resistance is due to an increased extrusion of PMEA and 6-thioinosine monophosphate from the cells that overproduce MRP5. In polarized Madin-Darby canine kidney II (MDCKII) cells transfected with an MRP5 cDNA construct, MRP5 is routed to the basolateral membrane and these cells transport S-(2,4-dinitrophenyl)glutathione and glutathione preferentially toward the basal compartment. Inhibitors of organic anion transport inhibit transport mediated by MRP5. We speculate that MRP5 might play a role in some cases of unexplained resistance to thiopurines in acute lymphoblastic leukemia and/or to antiretroviral nucleoside analogs in HIV-infected patients.
Cellular export of cyclic nucleotides has been observed in various tissues and may represent an elimination pathway for these
signaling molecules, in addition to degradation by phosphodiesterases. In the present study we provide evidence that this
export is mediated by the multidrug resistance protein isoform MRP5 (gene symbol ABCC5). The transport function of MRP5 was
studied in V79 hamster lung fibroblasts transfected with a humanMRP5 cDNA. An MRP5-specific antibody detected an overexpression of the glycoprotein of 185 ± 15 kDa in membranes from MRP5-transfected cells and a low basal expression of hamster Mrp5 in control membranes. ATP-dependent transport of 3′,5′-cyclic
GMP at a substrate concentration of 1 μmwas 4-fold higher in membrane vesicles fromMRP5-transfected cells than in control membranes. This transport was saturable with a K
m value of 2.1 μm. MRP5-mediated transport was also detected for 3′,5′-cyclic AMP at a lower affinity, with a K
mvalue of 379 μm. A potent inhibition of MRP5-mediated transport was observed by several compounds, known as phosphodiesterase modulators,
including trequinsin, with a K
i of 240 nm, and sildenafil, with a K
i value of 267 nm. Thus, cyclic nucleotides are physiological substrates for MRP5; moreover, MRP5 may represent a novel pharmacological target
for the enhancement of tissue levels of cGMP.
P-glycoprotein (Pgp) is a transmembrane protein conferring multidrug resistance to cells by extruding a variety of amphipathic cytotoxic agents using energy from ATP hydrolysis. The objective of this study was to understand how substrates affect the catalytic cycle of ATP hydrolysis by Pgp. The ATPase activity of purified and reconstituted recombinant human Pgp was measured using a continuous cycling assay. Pgp hydrolyzes ATP in the absence of drug at a basal rate of 0.5 micromol x min x mg(-1) with a K(m) for ATP of 0.33 mm. This basal rate can be either increased or decreased depending on the Pgp substrate used, without an effect on the K(m) for ATP or 8-azidoATP and K(i) for ADP, suggesting that substrates do not affect nucleotide binding to Pgp. Although inhibitors of Pgp activity, cyclosporin A, its analog PSC833, and rapamycin decrease the rate of ATP hydrolysis with respect to the basal rate, they do not completely inhibit the activity. Therefore, these drugs can be classified as substrates. Vanadate (Vi)-induced trapping of [alpha-(32)P]8-azidoADP was used to probe the effect of substrates on the transition state of the ATP hydrolysis reaction. The K(m) for [alpha-(32)P]8-azidoATP (20 microm) is decreased in the presence of Vi; however, it is not changed by drugs such as verapamil or cyclosporin A. Strikingly, the extent of Vi-induced [alpha-(32)P]8-azidoADP trapping correlates directly with the fold stimulation of ATPase activity at steady state. Furthermore, P(i) exhibits very low affinity for Pgp (K(i) approximately 30 mm for Vi-induced 8-azidoADP trapping). In aggregate, these data demonstrate that the release of Vi trapped [alpha-(32)P]8-azidoADP from Pgp is the rate-limiting step in the steady-state reaction. We suggest that substrates modulate the rate of ATPase activity of Pgp by controlling the rate of dissociation of ADP following ATP hydrolysis and that ADP release is the rate-limiting step in the normal catalytic cycle of Pgp.
P-glycoprotein (Pgp) is a plasma membrane protein whose overexpression confers multidrug resistance to tumor cells by extruding amphipathic natural product cytotoxic drugs using the energy of ATP. An elucidation of the catalytic cycle of Pgp would help design rational strategies to combat multidrug resistance and to further our understanding of the mechanism of ATP-binding cassette transporters. We have recently reported (Sauna, Z. E., and Ambudkar, S. V. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 2515-2520) that there are two independent ATP hydrolysis events in a single catalytic cycle of Pgp. In this study we exploit the vanadate (Vi)-induced transition state conformation of Pgp (Pgp.ADP.Vi) to address the question of what are the effects of ATP hydrolysis on the nucleotide-binding site. We find that at the end of the first hydrolysis event there is a drastic decrease in the affinity of nucleotide for Pgp coincident with decreased substrate binding. Release of occluded dinucleotide is adequate for the next hydrolysis event to occur but is not sufficient for the recovery of substrate binding. Whereas the two hydrolysis events have different functional outcomes vis à vis the substrate, they show comparable t(12) for both incorporation and release of nucleotide, and the affinities for [alpha-(32)P]8-azido-ATP during Vi-induced trapping are identical. In addition, the incorporation of [alpha-(32)P]8-azido-ADP in two ATP sites during both hydrolysis events is also similar. These data demonstrate that during individual hydrolysis events, the ATP sites are recruited in a random manner, and only one site is utilized at any given time because of the conformational change in the catalytic site that drastically reduces the affinity of the second ATP site for nucleotide binding. In aggregate, these findings provide an explanation for the alternate catalysis of ATP hydrolysis and offer a mechanistic framework to elucidate events at both the substrate- and nucleotide-binding sites in the catalytic cycle of Pgp.
The 190-kDa phosphoglycoprotein multidrug resistance protein 1 (MRP1) (ABCC1) confers resistance to a broad spectrum of anticancer drugs and also actively transports certain xenobiotics with reduced glutathione (GSH) (cotransport) as well as conjugated organic anions such as leukotriene C(4) (LTC(4)). In the present study, we have investigated a series of bioflavonoids for their ability to influence different aspects of MRP1 function. Most flavonoids inhibited MRP1-mediated LTC(4) transport in membrane vesicles and inhibition by several flavonoids was enhanced by GSH. Five of the flavonoids were competitive inhibitors of LTC(4) transport (K(i), 2.4-21 microM) in the following rank order of potency: kaempferol > apigenin (+ GSH) > quercetin > myricetin > naringenin (+ GSH). These flavonoids were less effective inhibitors of 17beta-estradiol 17beta-(D-glucuronide) transport. Moreover, their rank order of inhibitory potency for this substrate differed from that for LTC(4) transport inhibition but correlated with their relative lipophilicity. Several flavonoids, especially naringenin and apigenin, markedly stimulated GSH transport by MRP1, suggesting they may be cotransported with this tripeptide. Quercetin inhibited the ATPase activity of purified reconstituted MRP1 but stimulated vanadate-induced trapping of 8-azido-alpha-[(32)P]ADP by MRP1. In contrast, kaempferol and naringenin stimulated both MRP1 ATPase activity and trapping of ADP. In intact MRP1-overexpressing cells, quercetin reduced vincristine resistance from 8.9- to 2.2-fold, whereas kaempferol and naringenin had no effect. We conclude that dietary flavonoids may modulate the organic anion and GSH transport, ATPase, and/or drug resistance-conferring properties of MRP1. However, the activity profile of the flavonoids tested differed from one another, suggesting that at least some of these compounds may interact with different sites on the MRP1 molecule.
Human multidrug resistance protein 4 (MRP4) has recently been determined to confer resistance to the antiviral purine analog 9-(2-phosphonylmethoxyethyl)adenine and methotrexate. However, neither its substrate selectivity nor physiological functions have been determined. Here we report the results of investigations of the in vitro transport properties of MRP4 using membrane vesicles prepared from insect cells infected with MRP4 baculovirus. It is shown that expression of MRP4 is specifically associated with the MgATP-dependent transport of cGMP, cAMP, and estradiol 17-beta-D-glucuronide (E(2)17 beta G). cGMP, cAMP, and E(2)17 beta G are transported with K(m) and V(max) values of 9.7 +/- 2.3 microm and 2.0 +/- 0.3 pmol/mg/min, 44.5 +/- 5.8 microm and 4.1 +/- 0.4 pmol/mg/min, and 30.3 +/- 6.2 microm and 102 +/- 16 pmol/mg/min, respectively. Consistent with its ability to transport cyclic nucleotides, it is demonstrated that the MRP4 drug resistance profile extends to 6-mercaptopurine and 6-thioguanine, two anticancer purine analogs that are converted in the cell to nucleotide analogs. On the basis of its capacity to transport cyclic nucleotides and E(2)17 beta G, it is concluded that MRP4 may influence diverse cellular processes regulated by cAMP and cGMP and that its substrate range is distinct from that of any other characterized MRP family member.
Mercaptopurines have been used as anticancer agents for more than 40 years, and most acute lymphoblastic leukemias are treated with 6-mercaptopurine (6MP) or 6-thioguanine (TG). Overexpression of the two related multidrug resistance proteins MRP4 and MRP5 has been shown to confer some resistance against mercaptopurines, which has been attributed to extrusion of mercaptopurine metabolites by these transporters. We have analyzed the mercaptopurine metabolites formed in human embryonic kidney cells and determined which metabolites are extruded by MRP4 and MRP5. Incubation with 6MP led to the formation of thioinosine and thioxanthosine metabolites and we found that thio-IMP was transported by both MRP4 and MRP5; MRP5 showed the highest transport rate. In contrast, only MRP5 transported thioxanthosine monophosphate (tXMP). During incubation with TG, the monophosphorylated form of thioguanosine was transported by both MRP4 and MRP5; the highest transport rate was for MRP4. Similarly, only 6-methyl-thio-IMP was formed during incubation with 6-methyl mercaptopurine riboside. This compound was a substrate for both MRP4 and MRP5; MRP4 showed the highest transport rate. Our results show that all major thiopurine monophosphates important in the efficacy of mercaptopurine treatment are transported by MRP4 and MRP5, although the substrate specificity of the two transporters differs in detail.
Prostaglandins are involved in a wide variety of physiological and pathophysiological processes, but the mechanism of prostaglandin release from cells is not completely understood. Although poorly membrane permeable, prostaglandins are believed to exit cells by passive diffusion. We have investigated the interaction between prostaglandins and members of the ATP-binding cassette (ABC) transporter ABCC [multidrug resistance protein (MRP)] family of membrane export pumps. In inside-out membrane vesicles derived from insect cells or HEK293 cells, MRP4 catalyzed the time- and ATP-dependent uptake of prostaglandin E1 (PGE1) and PGE2. In contrast, MRP1, MRP2, MRP3, and MRP5 did not transport PGE1 or PGE2. The MRP4-mediated transport of PGE1 and PGE2 displayed saturation kinetics, with Km values of 2.1 and 3.4 microM, respectively. Further studies showed that PGF1alpha, PGF2alpha, PGA1, and thromboxane B2 were high-affinity inhibitors (and therefore presumably substrates) of MRP4. Furthermore, several nonsteroidal antiinflammatory drugs were potent inhibitors of MRP4 at concentrations that did not inhibit MRP1. In cells expressing the prostaglandin transporter PGT, the steady-state accumulation of PGE1 and PGE2 was reduced proportional to MRP4 expression. Inhibition of MRP4 by an MRP4-specific RNA interference construct or by indomethacin reversed this accumulation deficit. Together, these data suggest that MRP4 can release prostaglandins from cells, and that, in addition to inhibiting prostaglandin synthesis, some nonsteroidal antiinflammatory drugs might also act by inhibiting this release.
Recombinant nucleotide-binding domains (NBDs) from human multidrug resistance protein MRP1 were overexpressed in bacteria and purified to measure their direct interaction with high-affinity flavonoids, and to evaluate a potential correlation with inhibition of MRP1-mediated transport activity and reversion of cellular multidrug resistance. Among different classes of flavonoids, dehydrosilybin exhibited the highest affinity for both NBDs, the binding to N-terminal NBD1 being prevented by ATP. Dehydrosilybin increased vanadate-induced 8-N3-[alpha-32P]ADP trapping, indicating stimulation of ATPase activity. In contrast, dehydrosilybin strongly inhibited leukotriene C4 (LTC4) transport by membrane vesicles from MRP1-transfected cells, independently of reduced glutathione, and chemosensitized cell growth to vincristine. Hydrophobic C-isoprenylation of dehydrosilybin increased the binding affinity for NBD1, but outsite the ATP site, lowered the increase in vanadate-induced 8-N3-[alpha-32P]ADP trapping, weakened inhibition of LTC4 transport which became glutathione dependent, and induced some cross-resistance. The overall results indicate multiple binding sites for dehydrosilybin and its derivatives, on both cytosolic and transmembrane domains of MRP1.
Multidrug resistance protein 4 (MRP4/ABCC4), transports cyclic nucleoside monophosphates, nucleoside analog drugs, chemotherapeutic agents, and prostaglandins. In this study we characterize ATP hydrolysis by human MRP4 expressed in insect cells. MRP4 hydrolyzes ATP (Km, 0.62 mM), which is inhibited by orthovanadate and beryllium fluoride. However, unlike ATPase activity of P-glycoprotein, which is equally sensitive to both inhibitors, MRP4-ATPase is more sensitive to beryllium fluoride than to orthovanadate. 8-Azido[α-32P]ATP binds to MRP4 (concentration for half-maximal binding ∼3 μM) and is displaced by ATP or by its non-hydrolyzable analog AMPPNP (concentrations for half-maximal inhibition of 13.3 and 308 μM). MRP4 substrates, the prostaglandins E1 and E2, stimulate ATP hydrolysis 2- to 3-fold but do not affect the Km for ATP. Several other substrates, azidothymidme, 9-(2-phosphonylmethoxyethyl) adenine, and methotrexate do not stimulate ATP hydrolysis but inhibit prostaglandin E2-stimulated ATP hydrolysis. Although both post-hydrolysis transition states MRP4.8-azido[α- 32P]ABP·Vi and MRP4.8-azido[α-32P] ADP·beryllium fluoride can be generated, nucleotide trapping is ∼4-fold higher with beryllium fluoride. The divalent cations Mg2+ and Mn2+ support comparable levels of nucleotide binding, hydrolysis, and trapping. However, Co2+ increases 8-azido[α-32P]ATP binding and beryllium fluoride-induced 8-azido[α-32P]ADP trapping but does not support steady-state ATP hydrolysis. ADP inhibits basal and prostaglandin E2-stimulated ATP hydrolysis (concentrations for half-maximal inhibition 0.19 and 0.25 mM, respectively) and beryllium fluoride-induced 8-azido[α-32P]ADP trapping, whereas Pi has no effect up to 20 mM. In aggregate, our results demonstrate that MRP4 exhibits substrate-stimulated ATP hydrolysis, and we propose a kinetic scheme suggesting that ADP release from the post-hydrolysis transition state may be the rate-limiting step during the catalytic cycle.
The nature of cGMP transport in human erythrocytes, its relationship to glutathione conjugate transport, and possible mediation by multidrug resistance-associated proteins (MRPs) have been investigated. MRP1, MRP4 and MRP5 are detected in immunoblotting studies with erythrocytes. MRP1 and MRP5 are also detected in multidrug resistant COR-L23/R and MOR/R cells but at greatly reduced levels in the parent, drug sensitive COR-L23/P cells. MRP4 is detected in MOR/R but not COR-L23/R cells. Uptake of cGMP into inside-out membrane vesicles prepared by a spontaneous, one-step vesiculation process is shown to be by a low affinity system that accounts for more than 80% of the transport at all concentrations above 3 µm. This transport is reduced by MRP inhibitors and substrates including MK-571, methotrexate, estradiol 17-β-d-glucuronide, and S(2,4-dinitrophenyl)glutathione (DNP-SG) and also by glibenclamide and frusemide but not by the monoclonal Ig QCRL-3 that inhibits high-affinity transport of DNP-SG by MRP1. It is concluded that the cGMP exporter is distinct from MRP1 and has properties similar to those reported for MRP4. Furthermore the evidence suggests that the protein responsible for cGMP transport is the same as that mediating low-affinity DNP-SG transport in human erythrocytes.
Plant flavonoids show anti-inflammatory activity in vitro and in vivo. Although not fully understood, several action mechanisms are proposed to explain in vivo anti-inflammatory action. One of the important mechanisms is an inhibition of eicosanoid generating enzymes including phospholipase A2, cyclooxygenases, and lipoxygenases, thereby reducing the concentrations of prostanoids and leukotrienes. Recent studies have also shown that certain flavonoids, especially flavone derivatives, express their anti-inflammatory activity at least in part by modulation of proinflammatory gene expression such as cyclooxygenase-2, inducible nitric oxide synthase, and several pivotal cytokines. Due to these unique action mechanisms and significant in vivo activity, flavonoids are considered to be reasonable candidates for new anti-inflammatory drugs. To clearly establish the therapeutic value in inflammatory disorders, in vivo anti-inflammatory activity, and action mechanism of varieties of flavonoids need to be further elucidated. This review summarizes the effect of flavonoids on eicosanoid and nitric oxide generating enzymes and the effect on expression of proinflammatory genes. In vivo anti-inflammatory activity is also discussed. As natural modulators of proinflammatory gene expression, certain flavonoids have a potential for new anti-inflammatory agents.
Sarcolemmal vesicles prepared from rat heart exhibited ATP-dependent uptake of S-(2,4-dinitrophenyl)glutathione (DNP-SG), which obeyed Michaelis-Menten kinetics with an apparent Km of 21 microM for DNP-SG and a Vmax of 0.27 nmol.10 min-1.mg protein-1. Several model glutathione S-conjugates inhibited DNP-SG uptake, but leukotriene C4 inhibited uptake much more significantly even at lower concentrations (competitive inhibition, Ki = 1.5 microM). However, leukotrienes D4 and E4, which lack the gamma-glutamyl moiety, were less effective. The results suggest that the ATP-dependent transport system has a high affinity for leukotriene C4, and may be responsible for the translocation of this compound.
After almost 40 years of water fluoridation in the United States, its effect in the food chain is now being appreciated. Current surveys indicate significant increases in the F content of infant formulas, toddler cereals, fruit juices, and popular beverages, largely because fluoridated water is used in their processing. According to the best estimates, the daily total F intake of children from foods, beverages including water, and other sources such as unintentional ingestion of dentifrices containing F is on the rise, although it is generally within the currently accepted range for this age group. It is encouraging to note that appropriate steps are now being taken by some manufacturers of infant formulas to monitor F levels and keep them within an acceptable range. Because of the increasing contribution of dietary F to total F intake, dietary F should be included in any estimate of daily total F intake in children before F supplements are prescribed, whether the children live in communities with fluoridated or nonfluoridated water. To achieve this goal, it is essential to develop a generally accepted, sensitive method for the analysis of F in foods and beverages. This should help develop the bioavailability profiles for individual foods and beverages essential for accurate assessment of dietary F intake. Fluoridated salt, used in some European countries, appears to be the only food ingredient currently in use as an alternative to water fluoridation. Attempts to utilize staple foods and beverages as vehicles for systemic F delivery have generally failed because of the decreased bioavailability of F in such products, and because it is difficult to make them available to the general population and especially to the lower socioeconomic segments of the population.
Multidrug resistance is a major obstacle to cancer treatment. Using an expression cDNA library transfer approach to elucidating the molecular basis of non-P-glycoprotein-mediated multidrug resistance, we previously established that expression of multidrug resistance protein (MRP), an ATP-binding cassette superfamily transporter, confers multidrug resistance (G. D. Kruh et al., Cancer Res., 54: 1649-1652, 1994). In the present study, we generated NIH/3T3 MRP transfectants without using chemotherapeutic drugs to facilitate the pharmacological analysis of the MRP phenotype. MRP transfectants displayed increased resistance to several lipophilic drugs, including doxorubicin, daunorubicin, etoposide, actinomycin D, vincristine, and vinblastine. However, increased resistance was not observed for Taxol, a drug for which transfection of MDR1 confers high levels of resistance. Verapamil increased the sensitivity of MRP transfectants relative to control transfectants, but reversal was incomplete for doxorubicin and etoposide, the drugs for which MRP conferred the highest resistance levels. For the latter two drugs, MRP transfectants, which were approximately 8- and approximately 10-fold more sensitive than control cells in the absence of verapamil, exhibited 3.8- and 3.3-fold relative sensitization with 10 microM verapamil, respectively, but remained approximately 2 and approximately 3-fold more resistant than control cells. Analysis of drug kinetics using radiolabeled daunorubicin revealed decreased accumulation and increased efflux in MRP transfectants. Confocal microscopic analysis of intracellular daunorubicin in MRP transfectants was consistent with reduced intracellular drug concentrations, and also revealed an altered pattern of intracellular drug distribution characterized by the initial accumulation of drug in a perinuclear location, followed by the development of a punctate pattern of drug scattered throughout the cytoplasm. This pattern was suggestive of a process of drug sequestration, possibly followed by vesicle transport. Both increased drug efflux and perinuclear drug accumulation are consistent with the reported localization of MRP in plasma and cytosolic membranes (N. Krishnamachary and M. S. Center, Cancer Res., 53: 3658-3663, 1993; M. J. Flens et al., Cancer Res., 54: 4557-4563, 1994). These results thus indicate that the drug specificity of MRP is quite similar to that of MDR1, but also suggest potential differences in Taxol specificity and the level of verapamil sensitivity. In addition, these results indicate that MRP functions to extrude drug from the cell, but additionally suggest the intriguing possibility that drug sequestration contributes to drug resistance by protecting cellular targets and/or contributing to drug efflux.
Cell viability and in vitro cytotoxicity assay methods were developed using a combination of dyes, 4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate (WST-1), neutral red (NR) and crystal violet (CV), with HeLa cells as a bioindicatior. As WST-1 produces a highly water soluble and non-cytotoxic formazan dye, ti allows each assay to be carried out in one culture dish. The combined cell viability assay using WST-1, NR and CV gave an absorbance that correlated linearly with the number of cells over the range 1000 to 50,000 cells/well. The combined assay was applied to the evaluation of IC50 values for sodium dodecyl sulfate as a model toxicant, which yielded similar values to those obtained with each assay independently.
P-Glycoprotein (Pgp), a member of the adenosine triphosphate-binding cassette (ABC) superfamily of transporters, functions as an ATP-dependent drug efflux pump with broad specificity for chemically unrelated lipophilic compounds. This chapter discusses the Pgp ATPase activity that can be measured in the crude membranes of mouse (NIH 3T3) transfectants. The background activity of other ATPases can effectively be blocked with sodium azide. The inclusion of other inhibitors, such as ethylene glycol bis(β- aminoethyl ether)-N, N, N', N'-tetraacetic acid (EGTA) and ouabain does not have any deleterious effect on Pgp activity. NIH 3T3 transfectants selected at low drug concentration are also suitable for characterization of Pgp ATPase activity. NIH 3T3 cells are suitable for characterization of drug transport function and ATPase activity, as they offer an ideal stable expression system to assess the structure–function relationship of various regions in Pgp.
The main dietary sources of polyphenols are reviewed, and the daily intake is calculated for a given diet containing some common fruits, vegetables and beverages. Phenolic acids account for about one third of the total intake and flavonoids account for the remaining two thirds. The most abundant flavonoids in the diet are flavanols (catechins plus proanthocyanidins), anthocyanins and their oxidation products. The main polyphenol dietary sources are fruit and beverages (fruit juice, wine, tea, coffee, chocolate and beer) and, to a lesser extent vegetables, dry legumes and cereals. The total intake is approximately 1 g/d. Large uncertainties remain due to the lack of comprehensive data on the content of some of the main polyphenol classes in food. Bioavailability studies in humans are discussed. The maximum concentration in plasma rarely exceeds 1 microM after the consumption of 10-100 mg of a single phenolic compound. However, the total plasma phenol concentration is probably higher due to the presence of metabolites formed in the body's tissues or by the colonic microflora. These metabolites are still largely unknown and not accounted for. Both chemical and biochemical factors that affect the absorption and metabolism of polyphenols are reviewed, with particular emphasis on flavonoid glycosides. A better understanding of these factors is essential to explain the large variations in bioavailability observed among polyphenols and among individuals.
Despite accumulating evidence that multidrug resistance transporter proteins play a part in drug resistance in some clinical cancers, it remains unclear whether the relatively low levels of multidrug resistance transporter expression found in most untreated tumors could substantially affect their basal sensitivity to antineoplastic drugs. To shed light on this problem, the drug sensitivities of wild-type mouse cell lines were compared with those of lines in which the Mdr1a and Mdr1b genes encoding P-glycoprotein (P-gp) were inactivated and lines in which the Mrp1 gene was inactivated in addition to Mdr1a and Mdr1b. These models permit a clean dissection of the contribution of each transporter to drug resistance at expression levels similar to those in normal tissues and avoid complications that might arise from previous exposure of cell lines to drug selection. For substrate drugs, we found that these contributions can indeed be very substantial. Lines lacking functional P-gp were, on average, markedly more sensitive to paclitaxel (16-fold), anthracyclines (4-fold) and Vinca alkaloids (3-fold). Lines lacking both P-gp and Mrp1 were (compared with wild-type lines) hypersensitive to an even broader array of drugs, including epipodophyllotoxins (4-7-fold), anthracyclines (6-7-fold), camptothecins (3-fold), arsenite (4-fold) and Vinca alkaloids, especially vincristine (28-fold). Thus, even very low levels of P-gp and Mrp1 expression that may be difficult to detect in tumors could significantly affect their innate sensitivity to a wide range of clinically important substrate drugs. An implication is that the use of resistance reversal agents to sensitize drug-naive tumors may be appropriate in more cases than is presently appreciated.
Clotrimazole has been shown to have potent anti-malarial activity in vitro, one possible mechanism being inhibition of oxidized glutathione (GSSG) export from the infected human red blood cells or from the parasite itself. Efflux of GSSG from normal erythrocytes is mediated by a high affinity glutathione S-conjugate transporter. This paper shows that transport of the model substrate, 3 microm dinitrophenyl S-glutathione, across erythrocyte membranes is inhibited by multidrug resistance-associated protein 1 (MRP1)-specific antibody, QCRL-3, strongly suggesting that the high affinity transport is mediated by MRP1. The rates of transport observed with membrane vesicles prepared from erythrocytes or from multidrug resistant tumour cells show a similar pattern of responses to applied reduced glutathione, GSSG and MRP1 inhibitors (indomethacin, MK571) further supporting the conclusion that the high affinity transporter is MRP1. In both erythrocytes and MRP1-expressing tumour cells, MRP1-associated transport is inhibited by clotrimazole over the range 2-20 microm, and the inhibitory effect leads to increases in accumulation of MRP1 substrates, vincristine and calcein, and decreases in calcein efflux from intact MRP1-expressing human tumour cells. It also results in increased sensitivity to daunorubicin of the multidrug resistant cells, L23/R but not the sensitive parent L23/P cells. These results demonstrate that clotrimazole can inhibit the MRP1 which is present in human erythrocytes, an effect that may contribute to, though not fully account for, its anti-malarial action.
Chemotherapeutics are the most effective treatment for metastatic tumours. However, the ability of cancer cells to become simultaneously resistant to different drugs--a trait known as multidrug resistance--remains a significant impediment to successful chemotherapy. Three decades of multidrug-resistance research have identified a myriad of ways in which cancer cells can elude chemotherapy, and it has become apparent that resistance exists against every effective drug, even our newest agents. Therefore, the ability to predict and circumvent drug resistance is likely to improve chemotherapy.
The ATP-binding cassette (ABC) transporters are a family of large proteins in membranes and are able to transport a variety of compounds through membranes against steep concentration gradients at the cost of ATP hydrolysis. The available outline of the human genome contains 48 ABC genes; 16 of these have a known function and 14 are associated with a defined human disease. Major physiological functions of ABC transporters include the transport of lipids, bile salts, toxic compounds, and peptides for antigen presentation or other purposes. We review the functions of mammalian ABC transporters, emphasizing biochemical mechanisms and genetic defects. Our overview illustrates the importance of ABC transporters in human physiology, toxicology, pharmacology, and disease. We focus on three topics: (a) ABC transporters transporting drugs (xenotoxins) and drug conjugates. (b) Mammalian secretory epithelia using ABC transporters to excrete a large number of substances, sometimes against a steep concentration gradient. Several inborn errors in liver metabolism are due to mutations in one of the genes for these pumps; these are discussed. (c) A rapidly increasing number of ABC transporters are found to play a role in lipid transport. Defects in each of these transporters are involved in human inborn or acquired diseases.
To enable cell surface localization of the human multidrug resistance protein (MRP1, ABCC1) and to assess the role of the extracellular domains of this transporter, the FLAG epitope tag was introduced into different extracellular loops of the three membrane-spanning domains (MSDs) of the transporter. We constructed and expressed various partially and fully glycosylation-deficient, FLAG-tagged MRP1 proteins in a Vaccinia virus-based transient expression system, and the cell surface expression level of MRP1 on intact cells was followed by flow cytometry, using the FLAG tag specific monoclonal antibody M2. We also expressed the wild-type MRP1 protein and some of the FLAG-tagged mutants in stably transfected HEK293 cells, and followed the cell surface expression and the transport function of MRP1 both by monitoring the efflux of fluorescent substrate and by their ability to confer resistance to HEK293 transfectants to anticancer agents such as daunorubicin and etoposide. When we inserted the FLAG epitope in extracellular loops of the MSD1 or MSD3, the tag was accessible upon removal of N-glycosylation sites (N --> Q at positions 17, 23, and 1006, respectively), whereas the FLAG epitope placed in the MSD2 was not accessible even after removal of all three N-glycosylation sites, indicating that MSD2 region is deeply buried in the plasma membrane. However, all FLAG tagged MRP1 mutants were expressed at the cell surface to the same extent as the wild-type protein and also exhibited normal transport function. Our results demonstrate that the accessibility of the external FLAG epitope is strongly dependent on the position of the tag and the glycosylation state of the different FLAG-tagged MRP1s, and the conformation of extracellular loops in MSD1 and MDS3 does not appear to contribute to the functional status of MRP1.
The human MDR1 (ABCB1) gene product, P-glycoprotein (Pgp), functions as an ATP-dependent efflux pump for a variety of chemotherapeutic drugs. In this study, we assessed the role of conserved glutamate residues in the Walker B domain of the two ATP sites (E556 and E1201, respectively) during the catalytic cycle of human Pgp. The mutant Pgps (E556Q, E556A, E1201Q, E1201A, E556/1201Q, and E556/1201A) were characterized using a vaccinia virus based expression system. Although steady-state ATP hydrolysis and drug transport activities were abrogated in both E556Q and E1201Q mutant Pgps, [α-32P]-8-azidoADP was trapped in the presence of vanadate (Vi), and the release of trapped [α-32P]-8-azidoADP occurred to a similar extent as in wild-type Pgp. This indicates that these mutations do not affect either the first hydrolysis event or the ADP release step. Similar results were also obtained when Glu residues were replaced with Ala (E556A and E1201A). Following the first hydrolysis event and release of [α-32P]-8-azidoADP, both E556Q and E1201Q mutant Pgps failed to undergo another cycle of Vi-induced [α-32P]-8-azidoADP trapping. Interestingly, the double mutants E556/1201Q and E556/1201A trapped [α-32P]-8-azidoADP even in the absence of Vi, and the occluded nucleotide was not released after incubation at 37 °C for an extended period. In addition, the properties of transition state conformation of the double mutants generated in the absence of Vi were found to be similar to that of the wild-type protein trapped in the presence of Vi (Pgp·[α-32P]-8-azidoADP·Vi). Thus, in contrast to the single mutants, the double mutants appear to be defective in the ADP release step. In aggregate, these data suggest that E556 and E1201 residues in the Walker B domains may not be critical as catalytic carboxylates for the cleavage of the bond between the γ-P and the β-P of ATP during hydrolysis but are essential for the second ATP hydrolysis step and completion of the catalytic cycle.
The human multidrug resistance proteins MRP4 and MRP5 are organic anion transporters that have the unusual ability to transport cyclic nucleotides and some nucleoside monophosphate analogs. Base and nucleoside analogs used in the chemotherapy of cancer and viral infections are potential substrates. To assess the possible contribution of MRP4 and MRP5 to resistance against these drugs, we have investigated the transport mediated by MRP4 and MRP5. In cytotoxicity assays, MRP4 conferred resistance to the antiviral agent 9-(2-phosphonomethoxyethyl)adenine (PMEA) and high-performance liquid chromatography analysis showed that, like MRP5, MRP4 transported PMEA in an unmodified form. MRP4 also mediated substantial resistance against other acyclic nucleoside phosphonates, whereas MRP5 did not. Apart from low-level MRP4-mediated cladribine resistance, the cytotoxicity of clinically used anticancer nucleosides was not influenced by overexpression of MRP4 or MRP5. In contrast, MRP5 mediated efflux of the pyrimidine-based antiviral 2',3'-dideoxynucleoside 2',3'-didehydro-2',3'-dideoxythymidine 5'-monophosphate (d4TMP) and its phosphoramidate derivative alaninyl-d4TMP from cells loaded with the 2',3'-didehydro-2',3'-dideoxythymidine prodrugs cyclosaligenyl-d4TMP and aryloxyphosphoramidate d4TMP (So324), respectively. Moreover, only inside-out membrane vesicles derived from MRP5-overexpressing cells accumulated alaninyl-d4TMP. Cellular efflux and vesicular uptake studies were carried out to further compare transport mediated by MRP4 and MRP5 and showed that dipyridamole, dilazep, nitrobenzyl mercaptopurine riboside, sildenafil, trequinsin and MK571 inhibited MRP4 more than MRP5, whereas cyclic nucleotides and monophosphorylated nucleoside analogs were equally poor inhibitors of both pumps. These results strongly suggest that the affinity of MRP4 and MRP5 for nucleotide-based substrates is low.
The potency of flavonoids (isoflavones, flavones, and flavanones) to inhibit efflux of 2',7'-bis-(carboxypropyl)-5(6)-carboxyfluorescein (BCPCF) from human erythrocytes was investigated. Structure-activity relationship analysis showed that the strongest inhibitors were found among flavanones bearing a hydrophobic prenyl, geranyl, or lavandulyl group at position 8 (and hydroxyl groups at 5 and 7) in ring A. A prenyl group at position 5' or stilbene at positions 4'-5' in ring B further seemed to increase inhibitor potency. The most efficient flavanones, euchrestaflavanone A and sophoraflavanone H, were approximately 20 times more efficient than genistein, and induced 50% inhibition of BCPCF efflux (IC50) at 3 microM (60 min, 37 degrees C). This is comparable to IC50 of benzbromarone (4 microM) and lower than IC50 of indomethacin (10 microM), both known MRP1 (ABCC1) inhibitors. It is suggested that BCPCF efflux is mainly due to MRP1 activity. Our results indicate that flavonoid molecular structure provides a promising base for development of potent MRP1 inhibitors.
Quercetin is a typical flavonoid ubiquitously present in fruits and vegetables, and its antioxidant effect is implied to be helpful for human health. The bioavailability of quercetin glycosides should be clarified, because dietary quercetin is mostly present as its glycoside form. Although quercetin glycosides are subject to deglycosidation by enterobacteria for the absorption at large intestine, small intestine acts as an effective absorption site for glucose-bound glycosides (quercertin glucosides). This is because small intestinal cells possess a glucoside-hydrolyzing activity and their glucose transport system is capable of participating in the glucoside absorption. A study using a cultured cell model for intestinal absorption explains that the hydrolysis of the glucosides accelerates their absorption in the small intestine. Small intestine is also recognized as the site for metabolic conversion of quercetin and other flavonoids as it possesses enzymatic activity of glucuronidation and sulfation. Modulation of the intestinal absorption and metabolism may be beneficial for regulating the biological effects of dietary quercetin.
Multidrug transporters influence drug distribution in vivo and are often associated with tumour drug resistance. Here we show that plant-derived polyphenols that interact with P-glycoprotein can also modulate the activity of the recently discovered ABC transporter, breast cancer resistance protein (BCRP/ABCG2). In two separate BCRP-overexpressing cell lines, accumulation of the established BCRP substrates mitoxantrone and bodipy-FL-prazosin was significantly increased by the flavonoids silymarin, hesperetin, quercetin, and daidzein, and the stilbene resveratrol (each at 30 microM) as measured by flow cytometry, though there was no corresponding increase in the respective wild-type cell lines. These compounds also stimulated the vanadate-inhibitable ATPase activity in membranes prepared from bacteria (Lactococcus lactis) expressing BCRP. Given the high dietary intake of polyphenols, such interactions with BCRP, particularly in the intestines, may have important consequences in vivo for the distribution of these compounds as well as other BCRP substrates.
Breast cancer resistance protein (BCRP) is a newly identified ATP-binding cassette transporter, shown to confer multidrug resistance (MDR) to a number of important anticancer agents and play an important function in governing drug disposition. Flavonoids are a class of polyphenolic compounds widely present in foods and herbal products. The interactions of flavonoids with P-glycoprotein and multidrug resistance-associated protein 1 have been reported; however, their interaction with BCRP is unknown. Our objective was to evaluate the effects of 20 naturally occurring flavonoids on the cellular accumulation and cytotoxicity of mitoxantrone in both BCRP-overexpressing and BCRP-negative human cell lines. BCRP-overexpressing and BCRP-negative human breast cancer cells (MCF-7) and large cell lung carcinoma cells (NCI-H460) were used in these studies. Many of the tested flavonoids (50 microM) increased mitoxantrone accumulation in BCRP-overexpressing cells, completely reversing mitoxantrone resistance, with no effect on the corresponding BCRP-negative cells, indicating that these flavonoids are BCRP inhibitors. The effects of these flavonoids on the cellular accumulation and cytotoxicity of mitoxantrone were flavonoid concentration dependent, and significant changes were produced at concentrations lower than 10 microM for most of the flavonoids. Chrysin and biochanin A were the most potent BCRP inhibitors, producing significant increases in mitoxantrone accumulation at concentrations of 0.5 or 1.0 microM and in mitoxantrone cytotoxicity at a concentration of 2.5 microM. Flavonoid glycosides had no effects on the BCRP-mediated transport of mitoxantrone. The results obtained in this study could be clinically relevant in terms of both MDR reversal in cancer treatment and drug-flavonoid pharmacokinetic interactions.
Plant-derived flavonoids are inhibitors of various intracellular processes, notably phosphorylation pathways, and potential inhibitors of cellular autoimmunity. In this study, the inhibiting effects of various flavonoids on antigen-specific proliferation and interferon-gamma (IFN-gamma) production by human and murine autoreactive T cells were evaluated in vitro. T-cell responses were evaluated for the human autoantigen alpha B-crystallin, a candidate autoantigen in multiple sclerosis, and for the murine encephalitogen proteolipid protein peptide PLP (139-151). The flavones apigenin and luteolin were found to be strong inhibitors of both murine and human T-cell responses while fisitin, quercitin, morin and hesperitin, members of the subclasses of flavonoles and flavanones, were ineffective. Antigen-specific IFN-gamma production was reduced more effectively by flavones than T-cell proliferation, suggesting that the intracellular pathway for IFN-gamma production in T cells is particularly sensitive to flavone inhibition. These results indicate that flavones but not flavanoles or flavanones are effective inhibitors of the potentially pathogenic function of autoreactive T cells. The effects of flavones were the same for human and murine autoreactive T cells, stressing the usefulness of animal models of autoimmunity for further studies on the effects of flavonones on autoimmune diseases.
For analysis of multidrug resistance, a major barrier to effective cancer chemotherapy, we profiled mRNA expression of the 48 known human ABC transporters in 60 diverse cancer cell lines (the NCI-60) used by the National Cancer Institute to screen for anticancer activity. The use of real-time RT-PCR avoided artifacts commonly encountered with microarray technologies. By correlating the results with the growth inhibitory profiles of 1,429 candidate anticancer drugs tested against the cells, we identified which transporters are more likely than others to confer resistance to which agents. Unexpectedly, we also found and validated compounds whose activity is potentiated, rather than antagonized, by the MDR1 multidrug transporter. Such compounds may serve as leads for development.
Flavonoids are constituents of fruits, vegetables, and plant-derived beverages, as well as components in herbal-containing dietary supplements. The objective of this investigation was to characterize the effect of flavonoids on P-glycoprotein (P-gp)-mediated cellular efflux and to determine the molecular mechanism(s) of the flavonoid-drug interaction. Studies were conducted in the sensitive and multidrug resistant human breast cancer cell lines MCF-7 and MDA435/LCC6 and examined the effects of the flavonoids biochanin A, morin, phloretin, and silymarin on daunomycin (DNM) accumulation and doxorubicin cytotoxicity. The potential mechanism(s) involved in the interaction was evaluated by determining flavonoid effects on 1) P-gp ATPase activity, 2) [(3)H]azidopine photoaffinity labeling of P-gp, and 3) cellular P-gp levels. The flavonoids increased [(3)H]DNM accumulation in P-gp positive cells, but not P-gp negative cells, and these effects were both flavonoid concentration- and P-gp expression level-dependent. Biochanin A and silymarin potentiated doxorubicin cytotoxicity in P-gp positive cells. Biochanin A and phloretin stimulated, whereas morin and silymarin inhibited P-gp ATPase activity, confirming that these flavonoids interact with P-gp. Morin and silymarin significantly inhibited [(3)H]azidopine photoaffinity labeling of P-gp, suggesting a direct interaction with P-gp substrate binding. A 24-h preincubation with all flavonoids, followed by flavonoid removal, did not alter cellular P-gp level in P-gp positive cells. In conclusion, biochanin A, morin, phloretin, and silymarin all inhibited P-gp-mediated cellular efflux and the mechanism of the interaction involved, at least in part, a direct interaction. The findings of this study indicate a potential for significant flavonoid-drug interactions with P-gp substrates.
High dietary intake of fruits and vegetables is consistently associated with a reduced risk of common human cancers, including cancers of the lung, breast, prostate, and colon. It is unknown which bioactive compound or compounds in plant foods provide the chemoprotective effects. One class of compounds currently under investigation is flavonoids, a large group of compounds with similar structure, consisting of two phenolic benzene rings linked to a heterocyclic pyran or pyrone. Although there are numerous in vitro and animal model data suggesting that flavonoids influence important cellular and molecular mechanisms related to carcinogenesis, such as cell cycle control and apoptosis, there are limited data from human population studies. This article reviews data from four cohort studies and six case-control studies, which have examined associations of flavonoid intake with cancer risk. There is consistent evidence from these studies that flavonoids, especially quercetin, may reduce the risk of lung cancer. Further research using new dietary databases for food flavonoid content is needed to confirm these findings before specific public health recommendations about flavonoids can be formulated.
Flavonoids are food components that appear to have potential beneficial health effects. There is a range of in vitro studies supporting the anti-oxidant and anti-inflammatory properties of flavonoids. Previously, we demonstrated that in vitro flavonoids, including luteolin and apigenin, inhibit proliferation and IFN-gamma production by murine and human autoimmune T cells. In the present study, we examined the effects of oral flavonoids as well as of curcumin on autoimmune T cell reactivity in mice and on the course of experimental autoimmune encephalomyelitis (EAE), a model for multiple sclerosis. Continuous oral administration of flavonoids significantly affected antigen-specific proliferation and IFN-gamma production by lymph node-derived T cells following immunization with an EAE-inducing peptide. Both luteolin and apigenin suppress proliferative responses as they did in vitro, whereas IFN-gamma production on the other hand was enhanced. Other flavonoids exerted differential effects on proliferation and IFN-gamma production. The effects of flavonoids and curcumin on EAE were assessed using either passive transfer of autoimmune T cells or active disease induction. In passive EAE, flavonoids led to delayed recovery of clinical symptoms rather than to any reduction in disease. In active EAE, the effects were less pronounced but also, in this case, the flavonoid hesperitin delayed recovery. Oral curcumin had overall mild but beneficial effects. Our results indicate that oral flavonoids fail to beneficially influence the course of EAE in mice but, instead, suppress recovery from acute inflammatory damage.
Cystic fibrosis (CF) is caused by a mutation in the cystic fibrosis transmembrane conductance regulator (CFTR), a chloride channel expressed in epithelial cells. The effects of genistein and 4-phenylbutyrate (PBA) on CFTR were studied in three human airway epithelial cell lines expressing wild-type or DeltaF508 CFTR: Calu-3, CFSMEo-, and CFBE41o- cells. The cells were loaded with the fluorescent dye N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE) and chloride efflux was studied. Forskolin and 3-isobutyl-1-methylxanthine (IBMX) induced chloride efflux in Calu-3 cells but not in CF lines. Genistein (2.5-50 microM) alone was able to induce chloride efflux in all cell lines. Genistein did not enhance the effect of forskolin and IBMX. PBA had little or no effect on genistein-induced chloride efflux. The effect of genistein seen at low concentrations makes genistein interesting for possible pharmacological treatment of CF, since it is known that similar concentrations can be obtained in plasma by a soy-rich diet.
cGMP and glutathione-conjugate transport in human erythrocytes
- A Klokouzas
- Cp Wu
- Hw Van Veen
- Ma Barrand
- Sb Hladky
Klokouzas A, Wu CP, van Veen HW, Barrand MA, Hladky SB. cGMP and glutathione-conjugate
transport in human erythrocytes. Eur J Biochem 2003;270:3696–708. [PubMed: 12950253]
Multidrug-resistance protein 5 is a multispecific organic
- J Wijnholds
- Ca Mol
- L Van Deemter
- M De Haas
- Gl Scheffer
- F Baas
- Jh Beijnen
- Rj Scheper
- S Hatse
- De Clercq
- E Balzarini
- J Borst
Wijnholds J, Mol CA, van Deemter L, de Haas M, Scheffer GL, Baas F, Beijnen JH, Scheper RJ,
Hatse S, De Clercq E, Balzarini J, Borst P. Multidrug-resistance protein 5 is a multispecific organic
Dietary flavonoids and cancer risk: evidence from human population studies
- Ml Neuhouser
Neuhouser ML. Dietary flavonoids and cancer risk: evidence from human population studies. Nutr
Cancer 2004;50:1–7. [PubMed: 15572291]
Overexpression of multidrug resistance-associated protein (MRP) increases resistance to natural product drugs
- Ce Grant
- G Valdimarsson
- Dr Hipfner
- Kc Almquist
- Sp Cole
- Rg Deeley
Grant CE, Valdimarsson G, Hipfner DR, Almquist KC, Cole SP, Deeley RG. Overexpression of
multidrug resistance-associated protein (MRP) increases resistance to natural product drugs. Cancer
Res 1994;54:357–61. [PubMed: 8275468]
Mammalian ABC transporters in health and disease
- P Borst
- R Oude-Elferink
Hladky SB (2003) cGMP and glutathione-conjugate transport in human erythrocytes
- A Klokouzas
- Cp Wu
- Hw Van Veen
- Ma Barrand
Overexpression of multidrug resistance‐associated protein (MRP) increases resistance to natural product drugs
- Grant CE