Expression Of Adenosine Triphosphate-binding Cassette Transporters Research Articles

Casein Kinase 2α Augments Oxaliplatin Resistance in Colorectal Cancer Cells by Increasing ABCE1 Expression.

Antitumor drug resistance is a major hurdle in treating patients with malignant tumors. Casein kinase 2α (CK2α) expression is highly enhanced in oxaliplatin-resistant CRC cells. We investigated whether CK2α expression is associated with oxaliplatin resistance in CRC cells. To determine the effect of CK2α on drug resistance in CRC, we assessed the cell viability, adenosine triphosphate-binding cassette (ABC) transporter expression, apoptosis, and sphere formation according to CK2α expression in oxaliplatin-resistant CRC cells. CK2α expression was significantly increased in oxaliplatin-resistant CRC cells compared with that in wild-type CRC cells. In addition, the mRNA expression of ABC transporters, including ABCA12, ABCC2, and ABCE1, was significantly enhanced in oxaliplatin-resistant CRC cells, whereas this effect was blocked by the knockdown of CK2α. Furthermore, a cell viability test showed that oxaliplatin resistance was inhibited by decreasing CK2α expression, resulting in the induction of apoptosis and suppression of sphere formation. CK2α regulates cell survival, apoptosis, sphere formation, and drug resistance in oxaliplatin-resistant CRC cells by regulating ABC transporters. Therefore, targeting CK2α in drug-resistant CRC cells may be a novel strategy for treating patients with CRC.

Pxr- and Nrf2- mediated induction of ABC transporters by heavy metal ions in zebrafish embryos

Transcription factors including pregnane X receptor (Pxr) and nuclear factor-erythroid 2-related factor-2 (Nrf2) are important modulators of Adenosine triphosphate-binding cassette (ABC) transporters in mammalian cells. However, whether such modulation is conserved in zebrafish embryos remains largely unknown. In this manuscript, pxr- and nrf2-deficient models were constructed with CRISPR/Cas9 system, to evaluate the individual function of Pxr and Nrf2 in the regulation of ABC transporters and detoxification of heavy metal ions like Cd2+ and Ag+. As a result, both Cd2+ and Ag+ conferred extensive interactions with ABC transporters in wild type (WT) embryos: their accumulation and toxicity were affected by the activity of ABC transporters, and they significantly induced the mRNA expressions of ABC transporters. These induction effects were reduced by the mutation of pxr and nrf2, but elevations in the basal expression of ABC transporters compensated for the loss of their inducibility. This could be an explanation for remaining transporter function in both mutant models as well as the unaltered toxicity of metal ions in pxr-deficient embryos. However, mutation of nrf2 disrupted the production of glutathione (GSH), resulting in the enhanced toxicity of Cd2+/Ag+ in zebrafish embryos. In addition, elevated expressions of other transcription factors like aryl hydrocarbon receptor (ahr) 1b, peroxisome proliferator-activated receptor (ppar)-β, and nrf2 were found in pxr-deficient models without any treatment, while enhanced induction of ahr1b, ppar-β and pxr could only be seen in nrf2-deficient embryos after the treatment of metal ions, indicating different compensation phenomena for the absence of transcription factors. After all, pxr-deficient and nrf2-deficient zebrafish embryos are useful tools in the functional investigation of Pxr and Nrf2 in the early life stages of aquatic organisms. However, the compensatory mechanisms should be taken into consideration when interpreting the results and need in-depth investigations.

Oligomeric proanthocyanidins (OPCs) from grape seed extract suppress the activity of ABC transporters in overcoming chemoresistance in colorectal cancer cells.

Multidrug resistance is a major hindrance in managing cancer. By performing a series of experiments in chemoresistant colorectal cancer cell lines, we demonstrate that oligomeric proanthocyanidins (OPCs) from grape seed extracts can sensitize both acquired (HCT116-FOr cells) and innately chemoresistant (H716 cells) cancer cells to chemotherapeutic drugs, 5-fluorouracil (5FU) and oxaliplatin, by inhibiting adenosine triphosphate-binding cassette (ABC) transporter proteins. When combined with chemotherapeutic drugs, OPCs significantly inhibited growth of the chemoresistant cells (P < 0.05 to < 0.001) and decreased the expression of several key ABC transporters. Moreover, the activity of the ABC transporters was also significantly decreased by OPCs in the cell lines (P < 0.05). We further confirmed that co-treatment with OPCs sensitized the chemoresistant cells to 5FU and oxaliplatin, as observed by improvement in cell cycle arrest, double-strand breaks and p53 accumulation in these cells. In addition, we confirmed that co-administration of OPCs with chemotherapeutic drugs significantly decreased chemoresistant xenograft tumor growth in mice (P < 0.05). Together, our study illuminates the downregulation of multiple ABC transporters as a mechanism by which OPCs overcome chemoresistance in cancer cells and may serve as adjunctive treatments in patients with refractory colorectal cancer.

Altered Gene expression of ABC transporters, nuclear receptors and oxidative stress signaling in zebrafish embryos exposed to CdTe quantum dots

Adenosine triphosphate-binding cassette (ABC) transporters, including P-glycoprotein (Pgp) and multi-resistance associated proteins (Mrps), have been considered important participants in the self-protection of zebrafish embryos against environmental pollutants, but their possible involvement in the efflux and detoxification of quantum dots (QDs), as well as their regulation mechanism are currently unclear. In this work, gene expression alterations of ABC transporters, nuclear receptors, and oxidative stress signaling in zebrafish embryos after the treatment of mercaptopropionic acid (MPA)CdTe QDs and MPA-CdSCdTe QDs were investigated. It was observed that both QDs caused concentration-dependent delayed hatching effects and the subsequent induction of transporters like mrp1&2 in zebrafish embryos, indicating the protective role of corresponding proteins against CdTe QDs. Accompanying these alterations, expressions of nuclear receptors including the pregnane X receptor (pxr), aryl hydrocarbon receptor (ahr) 1b, and peroxisome proliferator-activated receptor (ppar)-β were induced by QDs in a concentration- and time-dependent manner. Moreover, elevated oxidative stress, reflected by the reduction of glutathione (GSH) level and superoxide dismutase (SOD) activities, as well as the dramatic induction of nuclear factor E2 related factor (nrf) 2, was also found. More importantly, alterations of pxr and nrf2 were more pronounced than that of mrps, and these receptors exhibited an excellent correlation with delayed hatching rate in the same embryos (R2 > 0.8). Results from this analysis demonstrated that the induction of mrp1 and mrp2 could be important components for the detoxification of QDs in zebrafish embryos. These transporters could be modulated by nuclear receptors and oxidative stress signaling. In addition, up-regulation of pxr and nrf2 could be developed as toxic biomarkers of CdTe QDs.

Transport mechanisms of hepatic uptake and bile excretion in clinical hepatobiliary scintigraphy with 99mTc-N-pyridoxyl-5-methyltryptophan

In clinical hepatobiliary scintigraphy, (99m)Tc-N-pyridoxyl-5-methyltryptophan ((99m)Tc-PMT) is an effective radiotracer among the (99m)Tc-pyridoxylaminates. However, the mechanisms of human hepatic uptake and bile excretion transport of (99m)Tc-PMT have not been determined. We thus investigated the transport mechanisms of human hepatic uptake and bile excretion in hepatobiliary scintigraphy with (99m)Tc-PMT. Four solute carrier (SLC) transporters involved in hepatic uptake were evaluated using human embryonic kidney (HEK) and HeLa cells with high expression of SLC transporters (organic anion transporting polypeptide (OATP)1B1, OATP1B3, OATP2B1, organic anion transporters (OAT)2 and organic cation transporters (OCT)1) after 5 min of (99m)Tc-PMT incubation. Metabolic analysis of (99m)Tc-PMT was performed using pooled human liver S9. Adenosine triphosphate (ATP)-binding cassette (ABC) transporters for bile excretion were examined using hepatic ABC transporter vesicles human expressing multiple drug resistance 1 (MDR1), multidrug resistance-associated protein 2 (MRP2), breast cancer resistance protein or bile salt export pump. (99m)Tc-PMT was incubated for 1, 3 and 5 min with ATP or adenosine monophosphate and these vesicles. SPECT scans were performed in normal and Eisai hyperbilirubinemic (EHBR) model rats, deficient in Mrp2 transporters, without and with verapamil (rat Mdr1 and human MDR1 inhibitor) after intravenous injection of (99m)Tc-PMT. Uptake of (99m)Tc-PMT in HEK293/OATP1B1 and HeLa/OATP1B3 was significantly higher than that in HEK293- and HeLa-mock cells. (99m)Tc-PMT was not metabolized in the human liver S9. In vesicles with high expression of ABC transporters, uptake of MDR1 or MRP2 was significantly higher at all incubation times. Bile excretion of (99m)Tc-PMT was also identified by comparison between normal and EHBR rats with and without verapamil on in-vivo imaging. Human hepatic uptake of (99m)Tc-PMT was transferred by OATP1B1 and OATP1B3, and excretion into bile canaliculi via MDR1 and MRP2. (99m)Tc-PMT hepatobiliary scintigraphy may be a useful ligand as a noninvasive method of visualizing and quantifying hepatobiliary transporter functionality, which could predict drug pharmacokinetics.

Expression of ABC Transporters is an Unfavorable Prognostic Factor in Laryngeal Squamous Cell Carcinoma

Two prominent and well-characterized representatives of adenosine triphosphate-binding cassette (ABC) transporter - breast cancer resistance protein (BCRP or ABCG2) and P-glycoprotein (P-gp or ABCB1) - are known to be membrane transporters associated with multidrug resistance. The aim of this study was to explore the correlation between ABC transporter expression and the clinicopathologic characteristics, proliferative index, and apoptotic index and their prognostic value in laryngeal squamous cell carcinoma (LSCC). Paraffin sections of 98 human LSCC specimens were investigated with immunohistochemical techniques. The relationship between ABCG2 and ABCB1 expression and clinicopathologic parameters, proliferative activities, and apoptotic activities and their prognostic value in patients' overall survival rate were subsequently analyzed. Immunohistochemical analysis revealed positive expression of ABCG2 and ABCB1 in 52.0% and 41.8% of patients, respectively. There was a positive correlation between ABCG2 expression and ABCB1 expression. The presence of these two proteins was significantly related to clinical stage, lymph node metastasis, and overall survival in LSCC. Interestingly, up-regulation of ABCG2 expression was found to be associated with increased proliferation, but that of ABCB1 was not. Up-regulation of both ABCG2 expression and ABCB1 expression was associated with decreased apoptosis. The results of this study revealed that the presence of ABCG2 and/or ABCB1 is predictive for malignant progression and is an independent prognostic factor in LSCC. The mechanism of ABC transporters may contribute to chemotherapy resistance by promoting proliferation and/or suppressing apoptosis.

PET and SPECT Radiotracers to Assess Function and Expression of ABC Transporters In Vivo

Adenosine triphosphate-binding cassette (ABC) transporters, such as P-glycoprotein (Pgp, ABCB1), breast cancer resistance protein (BCRP, ABCG2) and multidrug resistance-associated proteins (MRPs) are expressed in high concentrations at various physiological barriers (e.g. blood-brain barrier, blood-testis barrier, blood-tumor barrier), where they impede the tissue accumulation of various drugs by active efflux transport. Changes in ABC transporter expression and function are thought to be implicated in various diseases, such as cancer, epilepsy, Alzheimer's and Parkinson's disease. The availability of a non-invasive imaging method which allows for measuring ABC transporter function or expression in vivo would be of great clinical use in that it could facilitate the identification of those patients that would benefit from treatment with ABC transporter modulating drugs. To date three different kinds of imaging probes have been described to measure ABC transporters in vivo: i) radiolabelled transporter substrates ii) radiolabelled transporter inhibitors and iii) radiolabelled prodrugs which are enzymatically converted into transporter substrates in the organ of interest (e.g. brain). The design of new imaging probes to visualize efflux transporters is inter alia complicated by the overlapping substrate recognition pattern of different ABC transporter types. The present article will describe currently available ABC transporter radiotracers for positron emission tomography (PET) and single-photon emission computed tomography (SPECT) and critically discuss strengths and limitations of individual probes and their potential clinical applications.

Gene Expression Profiling of Adenosine Triphosphate-Binding Cassette Transporters in Response to K-ras Activation and Hypoxia in Human Pancreatic Cancer Cell Cultures

Pancreatic cancer (PC) is hypoxic and highly resistant to conventional chemotherapy. We sought to determine whether K-ras oncogene and/or hypoxia can induce expression of drug resistance-promoting adenosine triphosphate-binding cassette (ABC) transporters in human PC cell lines. Immortalized near-normal human pancreatic ductal epithelial(HPDE) cells, HPDE cells expressing K-rasG12V oncogene, and PCcell lines (MIA PaCa-2, PANC-1, BxPC-3) were subjected to hypoxia and examined for messenger RNA expression of 48 ABC transporters. Mutant K-ras activation and/or hypoxia of HPDE cells led to induction of various ABC transporters. In the case of PC cell lines, no clear correlation was found between expression of constitutively active K-ras and global ABC transporter expression. Moreover, hypoxic treatment of PC cell lines had different effects on ABC transporter expression.Importantly, PC cell lines did not express the multidrug resistance 1 ABC transporter, a major mechanism of drug resistance. However, multi drug resistance 1 expression in the cells was up-regulated in response to continuous exposure to low doses of vincristine, indicating that drug resistance could be induced. Expression of K-ras oncogene and hypoxia, as well as exposure to drugs, can contribute to drug resistance in PC cells.

Characterization of a side population of astrocytoma cells in response to temozolomide

Cancer progenitor-like cells isolated by Hoechst 33342 dye efflux (termed the "side population" [SP]) have been studied in a variety of cancers, including malignant brain tumors. In this study, the authors investigate the nature of the SP phenotype in 2 glioma cell lines, U87MG and T98G, and their response to temozolomide. The roles of several adenosine triphosphate-binding cassette (ABC) multidrug transporters expressed by SP cells, in particular ABCG2, are also examined. Using fluorescence-activated cell sorting, the cells were separated into SP and non-SP fractions and analyzed for progenitor cell-like properties with immunofluorescence staining, quantitative real-time polymerase chain reaction, and their ability to reform glioma mass in an immune-compromised mouse. The response of the SP cells to temozolomide was investigated at the cellular and molecular levels. Small interfering RNA knockdown was used to examine the specific role of the ABCG2 transporter, and the cells' tumorigenic potential was measured using the soft agar clonogenic assay. Side population cells are characterized by the presence of progenitor cell-like properties: increased expression of nestin, musashi-1, and ABCG2 were observed. In addition, only SP cells were able to reconstitute cellular heterogeneity; these cells were also more invasive than the non-SP cells, and possessed tumorigenic capacity. Temozolomide treatment increased the number of SP cells, and this corresponded to more progenitor-like cells, concurrent with elevated expression of several ABC transporters. Knockdown of ABCG2 transporters did not abrogate the SP cell response to temozolomide. Upregulation of several other ABC drug transporter genes is proposed to account for this chemoresistance.

Pediatric glioblastoma cell line shows different patterns of expression of transmembrane ABC transporters after in vitro exposure to vinblastine

Resistance to drug is a major cause of treatment failure in pediatric brain cancer. The multidrug resistance (MDR) phenotype can be mediated by the superfamily of adenosine triphosphate-binding cassette (ABC) transporters. The dynamics of expression of the MDR genes after exposure to chemotherapy, especially the comparison between pediatric brain tumors of different histology, is poorly described. To compare the expression profiles of the multidrug resistance genes ABCB1, ABCC1, and ABCG2 in different neuroepithelial pediatric brain tumor cell lines prior and following short-term culture with vinblastine. Immortalized lineages from pilocytic astrocytoma (R286), anaplasic astrocytoma (UW467), glioblastoma (SF188), and medulloblastoma (UW3) were exposed to vinblastine sulphate at different schedules (10 and 60 nM for 24 and 72 h). Relative amounts of mRNA expression were analyzed by real-time quantitative polymerase chain reaction. Protein expression was assessed by immunohistochemistry for ABCB1, ABCC1, and ABCG2. mRNA expression of ABCB1 increased together with augmenting concentration and time of exposure to vinblastine for R286, UW467, and UW3 cell lines. Interestingly, ABCB1 levels of expression diminished in SF188. Following chemotherapy, mRNA expression of ABCC1 decreased in all cell lines other than glioblastoma. ABCG2 expression was influenced by vinblastine only for UW3. The mRNA levels showed consistent association to protein expression in the selected sets of cell lines analyzed. The pediatric glioblastoma cell line SF188 shows different pattern of expression of multidrug resistance genes when exposed to vinblastine. These preliminary findings may be useful in determining novel strategies of treatment for neuroepithelial pediatric brain tumors.

ATP-binding cassette transporters in human heart failure

Adenosine triphosphate-binding cassette (ABC) transporters are involved in energy-dependent transport of substrates across biological membranes. We hypothesized that their expression is altered during human heart failure, suggesting a pathophysiologic basis. Messenger ribonucleic acid quantification of all known ABC transporters revealed multiple alterations in ABC transporter expression in failing human hearts (New York Heart Association classification III-IV) compared to nonfailing controls. These include a loss of ABCC7 chloride channels and an increased expression of the K(ATP) channel regulatory subunits ABCC8. Moreover, ABCG2, an efflux pump for xenobiotics/drugs, was expressed at much higher levels in failing hearts compared to nonfailing control hearts. ABCG2 was found in cardiac capillary endothelial cells and cardiomyocytes. Experiments in cells stably transfected with human ABCG2 revealed that the peroxisome proliferator-activated receptor-gamma agonist rosiglitazone was transported by ABCG2 but also inhibited the export of the prototypical ABCG2 substrate pheophorbide A (IC(50) 25 microM). These results suggest that altered ABC transporter expression in failing hearts might contribute to impaired channel conductance or might affect the cardiac disposition of drugs.

MDR1 and BCRP1 expression in leukemic progenitors correlates with chemotherapy response in acute myeloid leukemia

Overexpression of members of the adenosine triphosphate binding cassette (ABC) transporter superfamily has been implicated in multidrug resistance in cancer, but results in acute myeloid leukemia (AML) have been inconsistent. We investigated the expression and activity of ABC transporters in patient total blasts and subpopulations along the leukemic stem cell hierarchy. Using quantitative reverse transcriptase polymerase chain reaction, we measured expression of the ABC transporter superfamily in the blast cells from AML patients prior to chemotherapy. In addition, we measured ex vivo daunorubicin resistance of subpopulations with or without ABC inhibitors. In the total blasts, no consistent difference was observed in 18 patients achieving complete remission (CR) and 13 patients who were refractory to induction chemotherapy (NR). However, among the subpopulation of CD34(+)CD38(-) AML cells (candidate "leukemic stem cells"), elevated expression of MDR1 and/or BCRP1, two ABC transporters associated with drug resistance, was found in 8 of 10 NR patients as compared to 0 of 7 CR patients. No such association was observed in the more differentiated CD34(+)CD38(+) or CD34(-) subpopulations. There was no significant difference in MRP1 expression between CR and NR patient samples in any of the subpopulations examined. The increased expression of MDR1 and BCRP1 in leukemic cells correlated with increased cellular daunorubicin resistance, which could be reversed by the ABC transporter inhibitors verapamil and PSC-833. Expression of MDR1 and BCRP1 in leukemic stem cells correlates with chemotherapy response both at the cellular level and in AML patients.

Expression of Adenosine Triphosphate-Binding Cassette (ABC)??Drug Transporters in Peripheral??Blood Cells

Adenosine triphosphate-binding cassette (ABC)-type transport proteins were initially described for their ability to reduce intracellular concentrations of anticancer compounds, thereby conferring drug resistance. In recent years, expression of this type of proteins has also been reported in numerous cell types under physiological conditions; here, these transporters are often reported to alter systemic and local drug disposition (e.g. in the brain or the gastrointestinal tract). In this context, peripheral blood cells have also been found to express several ABC-type transporters. While erythrocytes mainly express multidrug resistance protein (MRP) 1, MRP4 and MRP5, which are discussed with regard to their involvement in glutathione homeostasis (MRP1) and in the efflux of cyclic nucleotides (MRP4 and MRP5), leukocytes also express P-glycoprotein and breast cancer resistance protein. In the latter cell types, the main function of efflux transporters may be protection against toxins, as these cells demonstrate a very high turnover rate. In platelets, only two ABC transporters have been described so far. Besides MRP1, platelets express relatively high amounts of MRP4 not only in the plasma membrane but also in the membrane of dense granules, suggesting relevance for mediator storage. In addition to its physiological function, ABC transporter expression in these structures can be of pharmacological relevance since all systemic drugs reach their targets via circulation, thereby enabling interaction of the therapeutic agent with peripheral blood cells. Moreover, both intended effects and unwanted side effects occur in peripheral blood cells, and intracellular micropharmacokinetics can be affected by these transport proteins. The present review summarises the data available on expression of ABC transport proteins in peripheral blood cells.

Reversal of ABC Transporter-Dependent Multidrug Resistance in Cancer

Multidrug resistance (MDR) remains the major cause of failure of chemotherapeutic treatment in cancer therapy. Thus, reversal of MDR in cancer is a desperately needed clinical requirement and a scientific challenge. The MDR phenotype can be mediated by adenosine triphosphate (ATP)-binding cassette (ABC) transporter molecules, which lower the intracellular accumulation of an entire panel of structurally and functionally unrelated anticancer drugs. Detailed knowledge of the transporter molecules, their sequence polymorphisms, their regulation at the transcriptional, translational, and post-translational levels, as well as their functional characteristics for substrate binding and transport, represent the basis for specific and, thus, successful intervention strategies for reversal of ABC transporter-dependent MDR in the clinic. Initial data obtained with third-generation inhibitors of ABC transporter-dependent MDR in clinical trials, and new developments with tools that interfere with expression of MDR genes, thereby leading to restored chemosensitization, are very promising attempts that provide grounds for cautious optimism. This article presents an overview of different strategies for overcoming ABC transporter-dependent MDR. Approaches targeting the function as well as the expression of MDR-related ABC transporters are summarized. Clinical trials evaluating the impact of compounds for reversal of ABC transporter-dependent MDR are discussed. The question remains, is reversal of MDR in cancer truly an achievable goal, given the different tumor entities and tumor heterogeneity, different tumor resistance profiles, and different therapeutic regimens? If so, MDR reversal will probably be successful only when tailored to an individual patient with an individual tumor, resistance profile and response prediction, i.e. individual prevention, circumvention, and reversal of MDR. Tailor-made MDR reversal strategies are the ultimate goal — and probably a realistic option.

Role of farnesoid X receptor in determining hepatic ABC transporter expression and liver injury in bile duct-ligated mice

Background & Aims : Cholestasis induces changes in hepatic adenosine triphosphate-binding cassette (ABC) transporter expression. We aimed to investigate the role of the nuclear bile acid receptor (farnesoid X receptor [FXR]) in mediating changes in ABC transporter expression and in determining liver injury. Methods : Hepatic ABC transporter (multidrug resistance-associated proteins [Mrp] 2–4 and bile salt export pump [Bsep]) expression and localization were studied in common bile duct-ligated (CBDL) FXR knockout (FXR −/−), wild-type (FXR +/+), and sham-operated mice. Serum alanine aminotransferase, alkaline phosphatase, bilirubin and bile acid levels, hepatic bile acid composition, and liver histology were investigated. Cholangiomanometry and bile duct morphometry were performed. Results : CBDL induced expression of Mrp 3 and Mrp 4 in FXR +/+ and even more in FXR −/−, whereas Mrp 2 expression remained unchanged. Bsep expression was maintained in CBDL FXR +/+ but remained undetectable in CBDL FXR −/−. Alanine aminotransferase levels and mortality rates did not differ between CBDL FXR +/+ and FXR −/−. CBDL increased biliary pressure and induced bile ductular proliferation and bile infarcts in FXR +/+, whereas FXR −/− had lower biliary pressures, less ductular proliferation, and developed disseminated liver cell necroses. Conclusions : Overexpression of Mrp 3 and Mrp 4 in CBDL mice is FXR independent and could play an important role in the adaptive hepatic ABC transporter response to cholestasis. Maintenance of Bsep expression strictly depends on FXR and is a critical determinant of the cholestatic phenotype. Lack of bile infarcts in CBDL FXR −/− suggests that development of bile infarcts is related to bile acid-dependent bile flow and biliary pressure. This information is relevant for the potential use of FXR modulators in the treatment of cholestatic liver diseases.