ABCA8: A potential therapeutic target in the treatment of colorectal cancer?

None.

Modulatory Activity of the Copper Chelate, Copper N-(2-Hydroxy Acetophenone) Glycinate, in ABC-transporter-expressing Cell Lines.

BACKGROUND/AIM: Metal-containing compounds (e.g., platinum complexes) belong to the standard armamentarium of cancer chemotherapy. Copper N-(2-hydroxy acetophenone) glycinate (CuNG) exerts anticancer activity in vitro and in vivo and modulates drug resistance related to glutathione or P-glycoprotein. The potential of CuNG to interact with ATP-binding cassette (ABC) transporters has not been fully explored yet. This study focused on the modulatory effects of CuNG on four ABC transporters (MRP1, MRP1, BCRP, and P-glycoprotein). MATERIALS AND METHODS: Cell viability, drug uptake and ABC transporter expression were measured by resazurin assays, flow cytometry, and ELISA in HL60AR, MDCKII-hBCRP, and Caco-2 cells. RESULTS: CuNG increased doxorubicin sensitivity of MRP1-over-expressing HL60AR with a similar efficacy as the control MRP1 inhibitor MK571. CuNG also increased MRP1's efflux activity. Comparable results were obtained with MDCKII cells over-expressing hBCRP. ELISA assays revealed that the expression of MRP1 in HL60AR cells and BCRP in MDCKII- cells was predominant but other ABC-transporters were also expressed at lower levels. Caco-2 cells expressed high levels of MRP2, but MRP1, BCRP, and P-glycoprotein were also expressed. In contrast to the two former cell lines, CuNG increased doxorubicin resistance and decreased efflux activity in Caco-2 cells. CONCLUSION: CuNG exerted different modulatory activities towards ABC-transporter-expressing cells. While CuNG-mediated ABC-transporter inhibition may improve tumor chemotherapy (like in HL60AR and MDCKII-hBCRP cells), CuNG-mediated enhanced ABC-transport (like in Caco-2 cells) may be a new strategy to ameliorate inflammatory diseases associated with decreased ABC-transporter expression such as ulcerative colitis.

Selective dysregulation of ABC transporters in methotrexate-resistant leukemia T-cells can confer cross-resistance to cytarabine, vincristine and dexamethasone, but not doxorubicin.

Pediatric acute lymphoblastic leukemia (pALL) includes 75 % of childhood leukemias, and methotrexate (MTX) is one of the most effective chemotherapy agents prescribed for pALL treatment. The aim of this study was to establish and characterize an MTX-resistant tumor cell model in order to study the mechanism contributing to drug sensitivity loss in pALL. Parental CCRF-CEM cells were treated with a gradual increasing concentration of MTX from 5 nM to 1.28 µM. The resistant subline was then characterized according to the cellular morphology, cellular growth curves and specific mRNA expression changes associated with drug resistance in ALL. Moreover, in vitro cytotoxicity assays were used to analyze cells relative responsiveness to a set of clinically used anti-ALL chemotherapy drugs. The morphological changes observed in the new R-CCRF-CEM/MVCD subline were associated with dysregulation of the EMT-related genes, Twist1 and CDH1. Cells demonstrated downregulation of ABCC1 and the overexpression of ABCA2, ABCA3, and ABCB1 membrane transporters. However, short treatment of the sensitive and parental cell line with MTX did not affect the expression profiles of the former ABC pumps. Moreover, R-CCRF-CEM/MVCD cells demonstrated cross-resistance to cytarabine (cytosine arabinoside, ara-C), vincristine, and dexamethasone, but not doxorubicin. The induced cross-resistance to specific chemotherapy drugs may possibly be attributed to selective dysregulation of the ABC transporters and EMT-related genes. These data may pave the way for the development of new cancer therapeutic strategies.

Pharmaceutical Excipients and Drug Metabolism: A Mini-Review.

Conclusions from previously reported articles have revealed that many commonly used pharmaceutical excipients, known to be pharmacologically inert, show effects on drug transporters and/or metabolic enzymes. Thus, the pharmacokinetics (absorption, distribution, metabolism and elimination) of active pharmaceutical ingredients are possibly altered because of their transport and metabolism modulation from the incorporated excipients. The aim of this review is to present studies on the interaction of various commonly-used excipients on pre-systemic metabolism by CYP450 enzymes. Excipients such as surfactants, polymers, fatty acids and solvents are discussed. Based on all the reported outcomes, the most potent inhibitors were found to be surfactants and the least effective were organic solvents. However, there are many factors that can influence the inhibition of CYP450, for instance type of excipient, concentration of excipient, type of CYP450 isoenzyme, incubation condition, etc. Such evidence will be very useful in dosage form design, so that the right formulation can be designed to maximize drug bioavailability, especially for poorly bioavailable drugs.

MRPA-independent mechanisms of antimony resistance in Leishmania infantum.

Control of both human and canine leishmaniasis is based on a very short list of chemotherapeutic agents, headed by antimonial derivatives (Sb). The utility of these molecules is severely threatened by high rates of drug resistance. The ABC transporter MRPA is one of the few key Sb resistance proteins described to date, whose role in detoxification has been thoroughly studied in Leishmania parasites. Nonetheless, its rapid amplification during drug selection complicates the discovery of other mechanisms potentially involved in Sb resistance. In this study, stepwise drug-resistance selection and next-generation sequencing were combined in the search for novel Sb-resistance mechanisms deployed by parasites when MRPA is abolished by targeted gene disruption. The gene mrpA is not essential in L. infantum, and its disruption leads to an Sb hypersensitive phenotype in both promastigotes and amastigotes. Five independent mrpA-/- mutants were selected for antimony resistance. These mutants displayed major changes in their ploidy, as well as extrachromosomal linear amplifications of the subtelomeric region of chromosome 23, which includes the genes coding for ABCC1 and ABCC2. Overexpression of ABCC2, but not of ABCC1, resulted in increased Sb tolerance in the mrpA-/- mutant. SNP analyses revealed three different heterozygous mutations in the gene coding for a serine acetyltransferase (SAT) involved in de novo cysteine synthesis in Leishmania. Overexpression of satQ390K, satG321R and satG325R variants led to a 2-3.2 -fold increase in Sb resistance in mrpA-/- parasites. Only satG321R and satG325R induced increased Sb resistance in wild-type parasites. These results reinforce and expand knowledge on the complex nature of Sb resistance in Leishmania parasites.

Vanillin confers antifungal drug synergism in Candida albicans by impeding CaCdr2p driven efflux.

AIM: Among the most common mechanisms of multidrug resistance (MDR) in prevalent human fungal pathogen, Candida albicans, overexpression of drug efflux pumps remains the predominant mechanism. Hence to inhibit efflux pumps and chemosensitize C. albicans against traditional antifungal drugs still represents an attractive approach. The present study aimed to analyze the effect of Vanillin (Van), a natural food flavoring agent, on drug efflux pump activity of Candida albicans. METHODS AND RESULTS: We observed that Van specifically inhibits Candida drug resistance protein 2 (CaCdr2p) activity belonging to ATP Binding Cassette (ABC) superfamily as revealed by abrogated rhodamine 6G efflux and nile red accumulation assay with CaCdr2p over expressing strain. Insight studies into the mechanisms suggested that abrogated efflux by CaCdr2p is due to competitive mode of inhibition by Van as depicted by Lineweaver-Burk plot. RT-PCR, western blot and confocal microscopy further unraveled that Van leads to reduced expression of CDR2 and CaCdr2p mislocalization respectively. Furthermore, Van sensitizes the azole sensitive and resistant clinical matched pair of isolates Gu4/Gu5 and led to abrogated rhodamine 6G efflux and depleted ergosterol. Furthermore, Van synergizes with membrane targeting drugs fluconazole and amphotericin B as their fractional inhibitory coefficient index was less than 0.5. CONCLUSION: Van being a potent inhibitor of CaCdr2p and chemosensitizing of drug resistant C. albicans warrants further studies to be exploited as effective antifungal agent.

Eucalyptal D Enhances the Antifungal Effect of Fluconazole on Fluconazole-Resistant Candida albicans by Competitively Inhibiting Efflux Pump.

The frequent emergence of azole-resistant strains has increasingly led azoles to fail in treating candidiasis. Combination with other drugs is a good option to effectively reduce or retard its incidence of resistance. Natural products are a promising synergist source to assist azoles in treating resistant candidiasis. Eucalyptal D (ED), a formyl-phloroglucinol meroterpenoid, is one of the natural synergists, which could significantly enhance the anticandidal activity of fluconazole (FLC) in treating FLC resistant C. albicans. The checkerboard microdilution assay showed their synergistic effect. The agar disk diffusion test illustrated the key role of ED in synergy. The rhodamine 6G (R6G) efflux assay reflected ED could reduce drug efflux, but quantitative reverse transcription PCR analysis revealed the upregulation of CDR1 and CDR2 genes in ED treating group. Efflux pump-deficient strains were hyper-susceptible to ED, thus ED was speculated to be the substrate of efflux pump Cdr1p and Cdr2p to competitively inhibit the excretion of FLC or R6G, which mainly contributed to its synergistic effect.

Interactions of Alectinib with Human ATP-Binding Cassette Drug Efflux Transporters and Cytochrome P450 Biotransformation Enzymes: Effect on Pharmacokinetic Multidrug Resistance.

Alectinib is a tyrosine kinase inhibitor currently used as a first-line treatment of anaplastic lymphoma kinase-positive metastatic nonsmall cell lung cancer (NSCLC). In the present work, we investigated possible interactions of this novel drug with ATP-binding cassette (ABC) drug efflux transporters and cytochrome P450 (P450) biotransformation enzymes that play significant roles in the phenomenon of multidrug resistance (MDR) of cancer cells as well as in pharmacokinetic drug-drug interactions. Using accumulation studies in Madin-Darby canine kidney subtype 2 (MDCKII) cells alectinib was identified as an inhibitor of ABCB1 and ABCG2 but not of ABCC1. In subsequent drug combination studies, we demonstrated the ability for alectinib to effectively overcome MDR in ABCB1- and ABCG2-overexpressing MDCKII and A431 cells. To describe the pharmacokinetic interaction profile of alectinib in a complete fashion, its possible inhibitory properties toward clinically relevant P450 enzymes (i.e., CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP3A4, or CYP3A5) were evaluated using human P450-expressing insect microsomes, revealing alectinib as a poor interactor. Advantageously for its use in pharmacotherapy, alectinib further exhibited negligible potential to cause any changes in expression of ABCB1, ABCG2, ABCC1, CYP1A2, CYP3A4, and CYP2B6 in intestine, liver, and NSCLC models. Our in vitro observations might serve as a valuable foundation for future in vivo studies that could support the rationale for our conclusions and possibly enable providing more efficient and safer therapy to many oncological patients.

Effect of photodynamic antimicrobial chemotherapy on Candida albicans in the presence of glucose.

BACKGROUND: Candida albicans is an opportunistic commensal microorganism, often associated with severe infections in immunosuppressed individuals. C. albicans has hexose transporters that may favor the intracellular accumulation of photosensitizer (PS). the aims of this study were to investigate the influence of glucose load on photodynamic antimicrobial chemotherapy (PACT); and the role that membrane transport system plays on this therapy in the presence of glucose. MATERIAL AND METHODS: Strains of C. albicans were selected: ATCC 10231, YEM 12, YEM 13, YEM 14 and YEM 15. All strains were grown aerobically on Sabouraud agar and incubated at 30 °C for 24 h. The strains were treated with and without glucose, and divided into Control (no treatment), LED light (660 nm, 166 mW/cm2), Photosensitizer (100 µM methylene blue) and PACT at 1, 3 and 6 min of irradiation groups. The colony forming units were counted and data submitted to statistical analysis (ANOVA) and Tukey's test. The concentration of methylene blue (MB) outside the yeast was measured by fluorescence spectroscopy. RESULTS: PACT inactivate C. albicans and the presence of glucose did not affect the killing effect for most strains. Only YEM12 was partially affected by its presence. Regarding efflux systems, ABC overexpressing strain showed a protective effect on the yeast cells. We observed that yeast with overexpression of major facilitator superfamily (MFS) membrane pore tended to accumulate more MB in its cytoplasm, whereas strains that overexpressed ABC pumps (ATP-binding cassette transporters) tended to decrease MB uptake and survive the photodynamic challenge. CONCLUSION: Presence of glucose showed a small effect on PACT . The accumulation of MB on yeast induces more photodynamic inactivation; however, the photodynamic efficacy depends on the type and characteristics of the microbial strain.

Modulation of the multixenobiotic resistance mechanism in Danio rerio hepatocyte culture (ZF-L) after exposure to glyphosate and Roundup®.

The presence of the transmembrane proteins of the ATP-binding cassette (ABC) family, which perform the efflux of several substances, contributes to the survival of aquatic organisms in a contaminated environmental. Those proteins provide a phenotype named the multixenobiotic resistance mechanism (MXR) by performing the efflux of a wide range of endogenous and exogenous compounds (ABCB) and biotransformation products and anionic compounds (ABCC). The aim of the present study was to evaluate the cellular defense pathway of an established culture from zebrafish hepatocytes (ZF-L) after 24 and 48 h of exposure to glyphosate and Original Roundup®, an herbicide used globally. Through abcb4, abcc1, abcc2 and abcc4 gene expression, ABCB and ABCC2 protein expression and ABC pump activity in ZF-L cells exposed to glyphosate and Roundup®. The results showed an increase in ABCB gene and protein expression; however, although ABCC2 showed an increase in gene expression, its protein expression was lower than in the control group. Regarding ABC activity, only exposure to Roundup® at the lowest concentration showed an increase at 48 h, but in the presence of inhibitors, both glyphosate and Roundup® appeared to modulate ABC activity, reducing its inhibition and returning activity to levels without inhibitor.

Role of drug transporters and heat shock proteins during ethanol exposure to human neural precursor cells and its lineages.

INTRODUCTION: Ethanol exposure to developing brain may alter the growth and differentiation of neurological cells resulting in unfavorable pathologies. Earlier studies have provided very limited mechanistic insights of cellular and molecular mechanisms which do not mimic with human situation due to varying cell types and poses potential challenges for investigation. Therefore, the present study was undertaken to evaluate the role of ABC transporters and heat shock proteins mediated response in human neural precursor cells (NPCs) and its lineages during proliferation and lineage differentiation against ethanol exposure. METHODS: Effect of ethanol exposure was examined for neuronal cell survival and variation in cellular phenotype during neurospheres development and lineage differentiation. Generation of reactive oxygen species, and variation in cell cycle was identified along with transcriptional profiling for pluripotent markers (Nestin, NCAM, Sox-2, and Notch-2), drug transporters (ABCB1 and ABCG2) and stress protein (HSP70) during ethanol exposure. RESULTS: ABC transporters as well as HSP70 mRNA expression was higher during proliferation as compared to differentiation with chronic ethanol (1 M) exposure (p < 0.01). Ethanol exposure resulted in higher variability in size and shape of developing neurospheres and decreased ability to form new neurosphere colonies. Significant changes were observed in dendrite development due to late ethanol exposure (p < 0.0001). CONCLUSION: The present study demonstrated significant role of ABC transporters and HSP70 proteins in providing defense against ethanol-induced damage in human neurological cells. However, the over-expression of ABC transporter and HSP-70 proteins during such pathological conditions do not provide complete defense and additional strategies are required to repair the damage.

Effect of ß-elemene on the kinetics of intracellular transport of d-luciferin potassium salt (ABC substrate) in doxorubicin-resistant breast cancer cells and the associated molecular mechanism.

In order to explore the mechanism of the reversing multidrug resistance (MDR) phenotypes by ß-elemene (ß-ELE) in doxorubicin (DOX)-resistant breast cancer cells (MCF-7/DOX), both the functionality and quantity of the ABC transporters in MCF-7/DOX were studied. Bioluminescence imaging (BLI) was used to study the efflux of d-luciferin potassium salt, the substrate of ATP-binding cassette transporters (ABC transporters), in MCF-7/DOX cells treated by ß-ELE. At the same time three major ABC transport proteins and genes-related MDR, P-glycoprotein (P-gp, ABCB1) and multidrug resistance-associated protein 1 (MRP, ABCC1) as well as breast cancer resistance protein (BCRP, ABCG2) were analyzed by q-PCR and Western blot. To investigate the efflux functionality of ABC transporters, MCF-7/DOXFluc cell line with stably-overexpressed luciferase was established. BLI was then used to real-time monitor the efflux kinetics of d-luciferin potassium salt before and after MCF-7/DOXFluc cells being treated with ß-ELE or not. The results showed that the efflux of d-luciferin potassium salt from MCF-7/DOXFluc was lessened when pretreated with ß-ELE, which means that ß-ELE may dampen the functionality of ABC transporters, thus decrease the efflux of d-fluorescein potassium or other chemotherapies which also serve as the substrates of ABC transporters. As the effect of ß-ELE on the expression of ABC transporters, the results of q-PCR and Western blot showed that gene and protein expression of ABC transporters such as P-gp, MRP, and BCRP were down-regulated after the treatment of ß-ELE. To verify the efficacy of ß-ELE on reversing MDR, MCF-7/DOX cells were treated with the combination of DOX and ß-ELE. MTT assay showed that ß-ELE increased the inhibitory effect of DOX on the proliferation of MCF-7/DOX, and the IC50 of the combination group was much lower than that of the single DOX or ß-ELE treatment. In all, ß-ELE may reverse MDR through the substrates of ABC transporters by two ways, to lessen the ABC protein efflux by weakening their functionality, or to reduce the quantity of ABC gene and protein expression.

Optimizing the flavanone core toward new selective nitrogen-containing modulators of ABC transporters.

AIM: Naringenin (1), isolated in large amount from the aerial parts of Euphorbia pedroi, was chemically derivatized to yield 18 imine derivatives (2-19) and three alkylated derivatives through a Mannich-type reaction (20-22) that were tested as multidrug resistance (MDR) reversers in cancer cells. Results/methodology: While hydrazone (2-4) and azine (5-13) derivatives showed an improvement in their MDR reversal activities against the breast cancer resistance protein, carbohydrazides 14-19 revealed an enhancement in MDR reversal activity toward the multidrug resistance protein 1. CONCLUSION: The observed activities, together with pharmacophoric analysis and molecular docking studies, identified the spatial orientation of the substituents as a key structural feature toward a possible mechanism by which naringenin derivatives may reverse MDR in cancer.

Characterisation of an ABC transporter of a resistant Candida glabrata clinical isolate.

BACKGROUND: Candida glabrata ranks second in epidemiological surveillance studies, and is considered one of the main human yeast pathogens. Treatment of Candida infections represents a contemporary public health problem due to the limited availability of an antifungal arsenal, toxicity effects and increasing cases of resistance. C. glabrata presents intrinsic fluconazole resistance and is a significant concern in clinical practice and in hospital environments. OBJECTIVE: The aim of this study was to characterise the azole resistance mechanism presented by a C. glabrata clinical isolate from a Brazilian university hospital. METHODS: Azole susceptibility assays, chemosensitisation, flow cytometry and mass spectrometry were performed. FINDINGS: Our study demonstrated extremely high resistance to all azoles tested: fluconazole, voriconazole, posaconazole and itraconazole. This isolate was chemosensitised by FK506, a classical inhibitor of ABC transporters related to azole resistance, and Rhodamine 6G extrusion was observed. A mass spectrometry assay confirmed the ABC protein identification suggesting the probable role of efflux pumps in this resistance phenotype. MAIN CONCLUSIONS: This study emphasizes the importance of ABC proteins and their relation to the resistance mechanism in hospital environments and they may be an important target for the development of compounds able to unsettle drug extrusion.

Histatin-5 induces the reversal of Pdr5p mediated fluconazole resistance in Saccharomyces cerevisae.

BACKGROUND: Candidiasis is a major opportunistic fungal infection in humans. The low number of antifungal drugs available to treat Candida infections and the increasing incidence of multidrug resistant (MDR) strains point to an urgent need of identifying new therapeutic options. The role of salivary components can provide insights for the development of new methodologies of control. OBJECTIVE: The aim of this study was to evaluate the ability of histatin-5, a constitutive immunological peptide present in saliva, in reversing fungal MDR phenotype, using a resistant Saccharomyces cerevisiae strain as model of study. RESULTS: A total of 2.5µg and 5µg of histatin-5 revealed to be able to chemosensitize (to revert antifungal resistance) a MDR strain to fluconazole impairing its intrinsic resistance. The presence of histatin-5 decreased the strain growth when associated to fluconazole, and also assisted in the retention of rhodamine 6G within cell cytoplasm. The ATPase activity of Pdr5p, an ABC efflux transporter, was significantly reduced up to 65% within physiological concentration of the peptide. CONCLUSION: Results revealed that histatin-5 is able to revert MDR phenotype and may be considered a potential alternative MDR inhibitor. Since Pdr5p is homologous to Candida albicans CaCdr1p and CaCdr2p, data obtained might be extrapolated to these transporters, inferring that associating fluconazole and histatin-5 may be a useful tool to circumvent failure treatments of infections caused by Candida MDR strains.

Cytotoxicity and Efflux Pump Inhibition Induced by Molybdenum Disulfide and Boron Nitride Nanomaterials with Sheetlike Structure.

Sheetlike molybdenum disulfide (MoS2) and boron nitride (BN) nanomaterials have attracted attention in the past few years due to their unique material properties. However, information on adverse effects and their underlying mechanisms for sheetlike MoS2 and BN nanomaterials is rare. In this study, cytotoxicities of sheetlike MoS2 and BN nanomaterials on human hepatoma HepG2 cells were systematically investigated at different toxic end points. Results showed that MoS2 and BN nanomaterials decreased cell viability at 30 µg/mL and induced adverse effects on intracellular ROS generation (≥2 µg/mL), mitochondrial depolarization (≥4 µg/mL), and membrane integrity (≥8 µg/mL for MoS2 and ≥2 µg/mL for BN). Furthermore, this study first found that low exposure concentrations (0.2-2 µg/mL) of MoS2 and BN nanomaterials could increase plasma membrane fluidity and inhibit transmembrane ATP binding cassette (ABC) efflux transporter activity, which make both nanomaterials act as a chemosensitizer (increasing arsenic toxicity). Damage to plasma membrane and release of soluble Mo or B species might be two reasons that both nanomaterials inhibit efflux pump activities. This study provides a systematic understanding of the cytotoxicity of sheetlike MoS2 and BN nanomaterials at different exposure levels, which is important for their safe use.

Serotonergic transmission at Merkel discs: modulation by exogenously applied chemical messengers and involvement of Ih currents.

The Merkel disc is a main type of tactile end organ consisting of Merkel cells and Aß-afferent endings that responds to tactile stimulation with slowly adapting type 1 (SA1) afferent impulses. Our recent study has shown that Merkel discs in whisker hair follicles are serotonergic synapses using endogenous serotonin to transmit tactile signals from Merkel cells to Aß-afferent endings. In this study, we hypothesize that tactile sensitivity of Merkel discs can be modulated by chemical messengers. We tested this hypothesis by determining whether and how SA1 responses of mouse whisker hair follicles may be affected by exogenously applied chemical messengers. We found that SA1 responses were potentiated by serotonin at low concentration (10 µM) but almost completely occluded by serotonin at high concentration (2 mM). In contrast, SA1 responses were not significantly affected by ATP and its metabolically stable analog α,ß-methylene-ATP, glutamate, γ-aminobutyric acid (GABA), and histamine. SA1 responses were also not affected by antagonists for P2X receptors, ionotropic glutamate receptors, and ionotropic GABA and glycine receptors. Whole-cell patch-clamp recordings reconfirm the presence of both ionotropic and metabotropic 5-HT receptors on afferent neurons and their terminals innervating whisker hair follicles. All whisker afferent neurons expressed hyperpolarization-activated inward currents (Ih ), which are potentiated by serotonin through the activation of metabotropic 5-HT receptors. Taken together, the findings substantiate the serotonergic mechanism of tactile transmission at Merkel discs and identify the involvement of Ih currents in postsynaptic excitatory actions of serotonin. In addition, the findings do not favor any significant involvement of ATP, glutamate, histamine, GABA, or glycine in tactile transmission at the Merkel discs of whisker hair follicles.

Impact of the induced organic anion transporter 1 (Oat1) renal expression by furosemide on the pharmacokinetics of organic anions.

AIM: Furosemide is a loop diuretic. Different authors demonstrated that continuous administration of furosemide modulates the expression of organic anion transporters. This study was undertaken to simultaneously evaluate the effects of furosemide pretreatment on organic anion transporter 1 (Oat1) and multidrug resistance protein 2 (Mrp2) renal expressions, on p-aminohippurate (PAH) pharmacokinetics and on renal and urinary PAH levels in rats. METHODS: Male Wistar rats were treated with furosemide (6 mg/100 g body weight per day, subcutaneously, 4 days) (treated group) or saline (control group). On the fifth day, PAH was administered as a bolus infusion in the femoral vein, and plasma samples were obtained from femoral artery at different time points. PAH levels in renal tissue and urine were also assessed. Renal Oat1 and Mrp2 expressions were evaluated by western blotting. RESULTS: Furosemide pretreatment increased both the expression of Oat1 and Mrp2. PAH plasma concentrations decreased following a biexponential function. The furosemide-treated group showed higher PAH plasma levels, a lower systemic clearance and elimination rate constant from the peripheral compartment, indicating that PAH renal elimination was decreased. PAH levels in renal tissue were significantly elevated and in urine appeared to be significantly lower as compared with control animals. CONCLUSIONS: Furosemide pretreatment caused a significant decrease of PAH renal elimination, despite Oat1 and Mrp2 augmented renal expression. The goal of the present study is the addition of important information in the wide gap of knowledge that exists about drug-drug interactions. Because of furosemide worldwide use, the data obtained are interesting and useful in terms of translation to clinical practice.

Extending the structure-activity relationship study of marine natural ningalin B analogues as P-glycoprotein inhibitors.

In the present study, a total of 25 novel ningalin B analogues were synthesized and evaluated for their P-gp modulating activity in a P-gp overexpressed breast cancer cell line LCC6MDR. Preliminary structure-activity study shows that A ring and its two methoxy groups are important pharmacophores for P-gp inhibiting activity. Among all derivatives, 23 is the most potent P-gp modulator with EC50 of 120-165 nM in reversing paclitaxel, DOX, vinblastine and vincristine resistance. It is relatively safe to use with selective index at least greater than 606 compared to verapamil. Mechanistic study demonstrates that compound 23 reverses P-gp mediated drug resistance by inhibiting transport activity of P-gp, thereby restoring intracellular drug accumulation. In summary, our study demonstrates that ningalin B analogue 23 is a non-cytotoxic and effective P-gp chemosensitizer that can be used in the future for reversing P-gp mediated clinical cancer drug resistance.