Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A subset of stem cells, termed "side population" (SP) cells, has been identified and characterized in several mammalian tissues and cell lines. However, SP cells have never been identified or isolated from gastrointestinal cancers. We used flow cytometry and the DNA-binding dye Hoechst 33342 to isolate SP cells from various human gastrointestinal system cancer cell lines. Fifteen of sixteen cancer cell lines from the gastrointestinal system contained 0.3%-2.2% SP cells. Next, we used an oligonucleotide microarray to analyze differentially expressed genes between SP and non-SP cells of hepatoma HuH7. The expression of GATA6, which is associated with embryonic development and hepatocytic differentiation, was significantly upregulated in HuH7 SP cells. The expression of ABCG2, ABCB1, and CEACAM6, which are associated with chemoresistance, was also significantly increased in SP cells. In addition, some epithelial markers and mesenchymal markers were overexpressed in SP cells. Reverse transcription-polymerase chain reaction and immunocytochemical staining validated these results and suggested a multilineage potential for HuH7 SP cells. In hepatoma HuH7 and colorectal SW480 cell lines, SP cells showed evidence for self-renewal, generating both SP and non-SP cells. Finally, chemoresistance to anticancer agents, including doxorubicin, 5-fluorouracil, and gemcitabine, were compared between HuH7 SP and non-SP cells using an ATP bioluminescence assay. The HuH7 SP cells expressed a higher resistance to doxorubicin, 5-fluorouracil, and gemcitabine compared with non-SP cells. These findings demonstrate that cancers of the gastrointestinal system do contain SP cells that show some characteristics of so-called stem cells.
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PMID:Characterization of a side population of cancer cells from human gastrointestinal system. 1623 20

The PLHC-1 hepatoma cell line derived from topminnow (Poeciliopsis lucida) is one of the most frequently used fish cell lines in aquatic ecotoxicology. These cells have been well characterized regarding the presence of phase I and phase II enzymes involved in the metabolism of xenobiotics. However, the presence of the ABC transport proteins possibly involved in the MultiXenobiotic Resistance (MXR) mechanism as phase III of cellular detoxification has never been described in the PLHC-1 cells. The main goal of this study was the detection and functional characterization of toxicologically relevant xenobiotic efflux transporters from ABCB and ABCC subfamily in the PLHC-1 cells. Using specific primer pairs two PCR products 1769 and 1023bp in length were successfully cloned and sequenced. Subsequent multiple alignment and phylogenetic analysis showed that these sequences share a high degree of homology with the P-glycoprotein (Pgp1; ABCB1) and the MRP3 (ABCC3). Functional experiments with fluorescent model substrates and specific inhibitors were used to verify that transport activities of Pgp- and MRP-related proteins are indeed present in PLHC-1 cells. Accumulation or efflux/retention rates of rhodamine 123, calcein-AM or monochlorbimane were time- and concentration-dependent. Cyclosporine A, MK571, verapamil, reversine 205, indomethacine and probenecid were used as specific inhibitors of Pgp1 and/or MRPs transport activities, resulting in a dose dependent inhibition of related transport activities in PLHC-1 cells. Similar to mammalian systems, the obtained IC(50) values were in the lower micromolar range. Taken together these data demonstrate that: (1) the PLHC-1 cells do express a functional MXR mechanism mediated by toxicologically relevant ABC efflux transporters; and (2) the presence of all three critical phases of cellular detoxification additionally affirms the PLHC-1 cells as a reliable in vitro model in aquatic toxicology.
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PMID:Detection and functional characterization of Pgp1 (ABCB1) and MRP3 (ABCC3) efflux transporters in the PLHC-1 fish hepatoma cell line. 1731 82

Frequently, primary hepatocytes are used as an in vitro model for the liver in vivo. However, the culture conditions reported vary considerably, with associated variability in performance. In this study, we characterized the differentiation character of primary human hepatocytes cultured using a highly defined, serum-free two-dimensional sandwich system, one that configures hepatocytes with collagen I as the substratum together with a dilute extracellular matrix (Matrigeltrade mark) overlay combined with a defined serum-free medium containing nanomolar levels of dexamethasone. Gap junctional communication, indicated by immunochemical detection of connexin 32 protein, was markedly enhanced in hepatocytes cultured in the Matrigel sandwich configuration. Whole genome expression profiling enabled direct comparison of liver tissues to hepatocytes and to the hepatoma-derived cell lines, HepG2 and Huh7. PANTHER database analyses were used to identify biological processes that were comparatively over-represented among probe sets expressed in the in vitro systems. The robustness of the primary hepatocyte cultures was reflected by the extent of unchanged expression character when compared directly to liver, with more than 77% of the probe sets unchanged in each of the over-represented categories, representing such genes as C/EBPalpha, HNF4alpha, CYP2D6, and ABCB1. In contrast, HepG2 and Huh7 cells were unchanged from the liver tissues for fewer than 48% and 55% of these probe sets, respectively. Further, hierarchical clustering of the hepatocytes, but not the cell lines, shifted from donor-specific to treatment-specific when the probe sets were filtered to focus on phenobarbital-inducible genes, indicative of the highly differentiated nature of the hepatocytes when cultured in a highly defined two-dimensional sandwich system.
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PMID:Gene expression profiling and differentiation assessment in primary human hepatocyte cultures, established hepatoma cell lines, and human liver tissues. 1751 62

Human cancers are characterized by a high degree of drug resistance. The multidrug resistance transporters MDR1-P-glycoprotein (ABCB1) and ABCC2 (MRP2) are expressed in a variety of human cancers, including hepatocellular carcinoma (HCC). The ABCC2 gene encodes a membrane protein involved in the ATP-dependent transport of conjugates of lipophilic substances. In this study we analyzed the effect of an ABCC2 antisense construct on the chemosensitization of HepG2 cells. Adenoviral vectors were constructed to allow an efficient expression of anti-ABCC2 antisense constructs. The effective target sequence comprised nucleotides 2543-2942 of the human ABCC2 cDNA. Adenoviral delivery of the ABCC2 antisense construct resulted in a reduced IC(50) for doxorubicin (12-fold), vincristine (50-fold), cisplatin (25-fold) and etoposide (VP-16) (25-fold). The adenoviral delivery of the ABCC2 antisense construct was so efficient that chemosensitization of HepG2 cells could even be demonstrated in mass cell cultures without a selection of transduced cells for single ABCC2 antisense-expressing HCC cell clones. After transfection of the ABCC2 antisense-expressing construct, HepG2 cells had significantly reduced ABCC2 mRNA and ABCC2 protein levels. Transduction of the ABCC2 antisense-expressing construct into HepG2 cells resulted in the accumulation of the high-affinity ABCC2 substrate Fluo-3. HepG2 tumors stably transfected with an anti-ABCC2 antisense construct regressed significantly in nude mice upon vincristine treatment. In addition, significant tumor regression was also observed when adenovirus-expressing anti-ABCC2 antisense construct was directly injected into HepG2 tumors in nude mice. Our study demonstrates the specific reversal of ABCC2-related drug resistance in adenovirus-transduced HepG2 cells and in HepG2 tumors in nude mice expressing this ABCC2 antisense construct.
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PMID:Reversal of drug resistance of hepatocellular carcinoma cells by adenoviral delivery of anti-ABCC2 antisense constructs. 1770 53

The development of the multidrug resistance (MDR) phenotype in mammals is often mediated by the overexpression of the P-glycoprotein1 (Pgp, ABCB1) or multidrug resistance-associated protein (MRP)-like ABC transport proteins. A similar phenomenon has also been observed and considered as an important part of the multixenobiotic resistance (MXR) defence system in aquatic organisms. We have recently demonstrated the presence of ABC transporters in the widely used in vitro fish model, the PLHC-1 hepatoma cell line. In the present study we were able to select a highly resistant PLHC-1 sub-clone (PLHC-1/dox) by culturing the wild-type cells in the presence of 1 microM doxorubicin. Using quantitative PCR a 42-fold higher expression of ABCB1 gene was determined in the PLHC-1/dox cells compared to non-selected wild-type cells (PLHC-1/wt). The efflux rates of model fluorescent Pgp1 substrates rhodamine 123 and calcein-AM were 3- to 4-fold higher in the PLHC-1/dox in comparison to the PLHC-1/wt cells. PLHC-1/dox were 45-fold more resistant to doxorubicin cytotoxicity than PLHC-1/wt. Similarly to mammalian cell lines, typical cross-resistance to cytotoxicity of other chemotherapeutics such as daunorubicin, vincristine, vinblastine, etoposide and colchicine, occurred. Furthermore, cyclosporine A, verapamil and PSC833, specific inhibitors of Pgp1 transport activity, completely reversed resistance of PLHC-1/dox cells to all tested drugs, resulting in EC50 values similar to the EC50 values found for PLHC-1/wt. In contrast, MK571, a specific inhibitor of MRP type of efflux transporters, sensitized PLHC-1/dox cells, neither to doxorubicin, nor to any other of the chemotherapeutics used in the study. These data demonstrate for the first time that a specific Pgp1-mediated doxorubicin resistance mechanism is present in the PLHC-1 fish hepatoma cell line. In addition, the fact that low micromolar concentrations of specific inhibitors may completely reverse a highly expressed doxorubicin resistance points to the fragility of Pgp1-mediated MXR defence mechanism in fish.
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PMID:Development and characterization of P-glycoprotein 1 (Pgp1, ABCB1)-mediated doxorubicin-resistant PLHC-1 hepatoma fish cell line. 1807 62

The ubiquitous presence of pharmaceuticals in aquatic systems is a challenging problem as their potential chronic effects on aquatic organisms remain largely unknown. The ATP-binding cassette (ABC) transport proteins contributing to the multidrug/multixenobiotic resistance (MDR/MXR) phenomenon seem to have an important role in the elimination of xenobiotics in aquatic organisms. Modulation of their efflux activities by contaminants may lead to substantial increase in intracellular accumulation and toxic effects of other xenobiotics. The aim of our work was to analyse a series of pharmaceuticals for their potential to modulate the activity of xenobiotic efflux transporters from the ABCB and ABCC sub-family in the Poeciliopsis lucida hepatoma cell (PLHC-1) fish cell line (PLHC-1/wt) and a doxorubicin (DOX) resistant PLHC-1 subclone (PLHC-1/dox) characterized by an elevated expression of the P-glycoprotein (ABCB1). Cellular accumulation of the model fluorescent substrates calcein-AM and rhodamine123 were used to determine an inhibitory effect on P-gp1 and/or MRP-like efflux transporters. 18 out of 33 tested pharmaceuticals showed MXR inhibitory activity with IC50 values occurring in the lower micromolar to millimolar range. Further, cytotoxic effects of pharmaceuticals were evaluated in PLHC-1/dox cells. Co-exposure of resistant cells to model P-gp1 inhibitor cyclosporine A (CyA) resulted in up to five times increased cytotoxicity of pharmaceuticals. In addition, some pharmaceuticals lead to a marked increase in cytotoxicity of doxorubicin, a model P-gp1 substrate. The modulation of toxicity by MDR inhibitors indicates their role in influencing cellular toxicity. In conclusion, the results of our study revealed significant inhibitory effects of environmentally relevant pharmaceuticals on P-gp1 and MRP-like transporters in fish. Our findings correspond well with data from mammalian systems indicating that the specificity and roles of the related efflux transporters may be similar in fish. Furthermore, due to the presence of active and inducible ABC transport proteins, the PLHC-1 cells appear to be a reliable in vitro system for the investigation of MDR/MXR mechanisms in fish.
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PMID:Human pharmaceuticals modulate P-gp1 (ABCB1) transport activity in the fish cell line PLHC-1. 1895 Aug 75

The mechanisms underlying atorvastatin supression of ABCB1 gene expression, at transcriptional and post-transcriptional levels of ABCB1 gene in HepG2 (human hepatocellular carcinoma) cells were investigated. Quantitative real-time PCR was used to measure mRNA levels, as well as to estimate the half-life of ABCB1 mRNA. Western blotting analysis was performed in order to measure protein levels of ABCB1. Electrophoretic mobility shift assay (EMSA) was used to evaluate interactions between protein(s) and ABCB1 promoter region. Exposure to atorvastatin for 24h resulted in a dose-dependent decrease of ABCB1 mRNA and protein levels, which was not abolished by addition of farnesyl or geranylgeranyl pyrophosphate. After removing fetal bovine serum from the media, however, ABCB1 expression was decreased by 2-fold in either HepG2 cells treated and non-treated with atorvastatin. Addition of cholesterol to serum free media abolished this latter effect on ABCB1 mRNA levels. In EMSA using a 5'-end-labeled 241bp ABCB1 promoter DNA fragment (-198 to +43) as probe, the binding of the proteins to the probe was reduced by NF-Y, but not changed by NFkappaB, AP-1, and SP1. However, the NF-Y binding activity was similar in control and atorvastatin-treated cells. mRNA stability studies revealed that ABCB1 mRNA degradation was increased in 1, 10 and 20muM atorvastatin-treated versus control cells (half-lives of 2h versus 7h). Therefore, evidence is provided that decreased mRNA stability by atorvastatin treatment may explain the decrease in ABCB1 transcript levels.
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PMID:Decreased ABCB1 mRNA expression induced by atorvastatin results from enhanced mRNA degradation in HepG2 cells. 1939 20

ABCB1 (P-glycoprotein) is an efflux transporter that limits the cellular uptake levels of various drugs in intestine, brain, and other tissues. The expression of human ABCB1 has recently been reported to be under the control of nuclear receptor NR1I subfamily members, pregnane X receptor (PXR, NR1I2) and constitutive androstane receptor (CAR, NR1I3). Here, we have investigated the involvement of another NR1I member, vitamin D receptor (VDR, NR1I1), in ABCB1 expression. In the human colorectal adenocarcinoma cell line LS174T, which abundantly expresses VDR, both 1alpha,25-dihydroxyvitamin D(3) (1,25-VD3) and lithocholic acid (LCA) increased ABCB1 mRNA levels. Reporter gene assays in LS174T cells with constructs containing various lengths of the ABCB1 regulatory region revealed that the region containing multiple nuclear receptor binding motifs located at -7.8 kilobases [termed nuclear receptor-responsive module (NURREM)], to which PXR and CAR also bind, is essential for the VDR-mediated ABCB1 transactivation. Further reporter assays with constructs containing truncated NURREM and gel shift assays suggested simultaneous binding of multiple VDR/retinoid X receptor alpha heterodimers to NURREM. Furthermore, knockdown of VDR expression in LS174T cells blocked the LCA- and the 1,25-VD3-induced transcription of ABCB1 reporter genes. In human hepatoma HepG2 cells, in contrast with LS174T cells, 1,25-VD3 activated the ABCB1 transcription only in the presence of ectopically expressed VDR. These results suggest that the NR1I subfamily members regulate the ABCB1 expression sharing the binding sites within NURREM and that the physiologically produced LCA and 1,25-VD3 may modulate the ABCB1 expression in human intestines, possibly associated with interindividual variations of ABCB1 expression.
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PMID:Involvement of Vitamin D receptor in the intestinal induction of human ABCB1. 1946 Sep 46

Both our previous study and other reports have suggested that CD133, originally classified as a hematopoietic stem cell marker, could be used for enrichment of cancer stem cells (CSCs) in human hepatocellular carcinoma (HCC). It was also noted that not all of CD133(+) cells were representative of CSCs. Further identification and characterization of CSCs or tumor-initiating cells in HCC are necessary to better understand hepatocarcinogenesis. In present study, we demonstrated that CSC phenotype could be precisely defined by co-expression of CD133 and CD44 cell surface markers. CD133(+)CD44(+) HCC cells showed stem cell properties, including extensive proliferation, self-renewal, and differentiation into the bulk of cancer cells. In vivo xenograft experiments revealed that, actually, the highly tumorigenic capacity of CD133(+) cells as previously described was primarily attributed to CD133(+)CD44(+) cell subpopulation, instead of their CD133(+)CD44(-) counterparts. Moreover, cells double-positive for CD133 and CD44 exhibited preferential expression of some stem cell-associated genes and were more resistant to chemotherapeutic agents due to the upregulation of ATP-binding cassette (ABC) superfamily transporters, including ABCB1, ABCC1, and ABCG2, further supporting these cells as HCC cell origin. Our findings suggest that CD133(+)CD44(+) cells might represent true cancer stem/progenitor cells in HCC, which could allow a better understanding of HCC initiation and progression, as well as establish a precise target for the development of more effective therapies.
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PMID:Cancer stem/progenitor cells are highly enriched in CD133+CD44+ population in hepatocellular carcinoma. 1971 46

Sorafenib is a second-generation, orally active multikinase inhibitor that is approved for the treatment of patients with advanced renal cell carcinoma and patients with unresectable hepatocellular carcinoma. We studied active transport of sorafenib in MDCK-II cells expressing human P-glycoprotein (P-gp/ABCB1) or ABCG2 (breast cancer resistance protein) or murine Abcg2. Sorafenib was moderately transported by P-gp and more efficiently by ABCG2 and Abcg2. Because sorafenib is taken orally, we orally administered sorafenib to wild-type, Abcb1a/1b(-/-), Abcg2(-/-), and Abcb1a/1b;Abcg2(-/-) mice, completely lacking functional Abcb1a/1b, Abcg2, or both, respectively, and we studied plasma pharmacokinetics and brain accumulation. The systemic exposure on oral administration was not different among all strains. However, brain accumulation was 4.3-fold increased in Abcg2(-/-) mice and 9.3-fold increased in Abcb1a/1b;Abcg2(-/-) mice. Moreover, when wild-type mice were treated with sorafenib in combination with the dual P-gp and ABCG2 inhibitor elacridar, brain accumulation was similar to that observed for Abcb1a/1b;Abcg2(-/-) mice. These results show that the brain accumulation of sorafenib is primarily restricted by ABCG2. This contrasts with previous studies using shared ABCG2 and P-gp substrates, which all suggested that P-gp dominates at the blood-brain barrier, and that an effect of ABCG2 is only evident when both transporters are absent. Interestingly, for sorafenib, it is the other way around, that is, ABCG2, and not P-gp, plays the dominant role in restricting its brain accumulation. Clinically, our findings may be relevant for the treatment of renal cell carcinoma patients with central nervous system relapses, as a dual ABCG2 and P-gp inhibitor might improve the central nervous system entry and thereby the therapeutic efficacy of sorafenib.
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PMID:Breast cancer resistance protein and P-glycoprotein limit sorafenib brain accumulation. 2010


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