Multidrug Resistance 1 Promoter Research Articles

Metformin alleviates adriamycin resistance of osteosarcoma by declining YY1 to inhibit MDR1 transcriptional activity

Chemotherapy resistance hinders the successful treatment of osteosarcoma (OS) to some extent. Previous studies have confirmed that metformin (Met) enhances apoptosis induced by chemotherapeutic drugs, but the underlying mechanism remains unclear. To establish adriamycin (ADM)-resistant MG-63 (MG-63/ADM) cells, the dosage of ADM was progressively increased. The results of qRT-PCR and Western blotting demonstrated that the expression level of Yin Yang 1 (YY1) and multi-drug resistance-1 (MDR1) in MG-63/ADM cells were remarkably increased compared with those in MG-63 cells. Met dramatically enhanced ADM cytotoxicity and accelerated apoptosis of MG-63/ADM cells. Moreover, Met suppressed the expressions of YY1 and MDR1 in MG-63/ADM cells. YY1 promoted its transcriptional expression by directly binding to the MDR1 promoter. Furthermore, the effects of Met on ADM sensitivity in MG-63/ADM cells was reversed due to overexpression of YY1 or MDR1. Collectively, these findings suggested that Met inhibited YY1/MDR1 pathway to reverse ADM resistance in OS, providing a new insight into the mechanism of Met in ADM resistance of OS.

Does MDR1 Promoter Methylation Affect Temozolomide Resistance? A Clinical Study in Patients with Glioblastoma

Purpose: The multidrug resistance 1 (MDR1) gene expression and its epigenetic status may be an important factor in the chemotherapeutic resistance of glioblastoma (GB). The aim of this study was to analyze the effect of the MDR1 promoter methylation status on GB tumor tissue related with patient survival, chemotherapy resistance, and recurrence of the disease. Materials and methods: Thirty-six patients underwent surgery for GB at the Neurosurgery Department of Ankara University School of Medicine. The patients’ clinical information and the MDR1 methylation status of the tumor tissues was compared to determine the effects on patient survival, chemotherapy resistance, and tumor recurrence. Results: Patients with MDR1 methylated GB had statistically significantly (p<0.001) shorter survival times. Early recurrence was detected in 25% of the patients with unmethylated tumor tissues and in 39.3% with hemimethylated tumor tissues. Conclusion: Instead of using the standard chemotherapeutics in all the patients with GB, tissue-specific medications must be chosen while taking into consideration the epigenomic characteristics and expression status of the tumor because of the genetic heterogeneity of GB. This is the first study to show the association between MDR1 promoter methylation and the clinical data of GB in the literature.

Visfatin mediates doxorubicin resistance in human colorectal cancer cells via up regulation of multidrug resistance 1 (MDR1).

Colorectal cancer (CRC) is one of the prevalent and deadly cancers worldwide. Chemotherapy resistance is one of the most challenging problems for CRC and other cancer treatments. Recent studies indicated that increasing levels of visfatin are correlated with worse clinical prognosis of CRC patients, while the effects and mechanisms of visfatin on progression of CRC remain unclear. Our present study established doxorubicin (Dox)-resistant CRC HCT-116 and SW480 cells (named HCT-116 Dox/R and SW480 Dox/R). The expression of visfatin, while not IL-6, IL-8, or TGF-β, in CRC Dox-resistant cells was significantly greater than that in their parental cells, while knockdown of visfatin by its specific siRNAs can elevate Dox sensitivity of CRC-resistant cells. In addition, si-visfatin can significantly down regulate the expression of multidrug resistance 1 (MDR1), while not multidrug resistance-associated protein 1 or lung resistance-related protein, in both HCT-116 Dox/R and SW480 Dox/R cells. Visfatin can regulate the transcription of MDR1 via modulating its promoter activities. Si-visfatin can also decrease the activation and nuclear localization of p65, one of the most important transcription factors for the expression of MDR1. Chromatin immunoprecipitation (ChIP) indicated that si-visfatin can suppress the binding between p65 and MDR1 promoter. Collectively, our present study revealed that visfatin mediates the Dox resistance of CRC cells via up regulation of MDR1. It indicated that targeted inhibition of visfatin might be helpful for overcoming Dox resistance of CRC therapy.

Pygopus2 inhibits the efficacy of paclitaxel-induced apoptosis and induces multidrug resistance in human glioma cells.

Anti-microtubule drugs, such as paclitaxel (PTX), are extensively used for the treatment of numerous cancers. However, growing evidence has shown that PTX resistance, either intrinsic or acquired, frequently occurs in patients and results in the failure of treatment, contributing to the high cancer mortality rate. Therefore, it is necessary to identify the genes or pathways involved in anti-microtubule drug resistance for future successful treatment of cancers. Pygopus2 (Pygo2), which contains a Zn-coordinated plant homeodomain (PHD) finger domain, is critical for β-catenin-dependent transcriptional switches in normal and malignant tissues and is over-expressed in various cancers, including human brain glioma. In this study, we report that over-expression of Pygo2 inhibited the efficacy of PTX and contributed to cell multidrug resistance in two different ways. First, over-expression of Pygo2 inhibited the PTX-induced phosphorylation of B-cell lymphoma 2 (Bcl-2), suppressing the proteolytic cleavage of procaspase-8/9 and further inhibiting the activation of caspase-3, which also inhibits the activation of the JNK/SAPK pathway, ultimately inhibiting cell apoptosis. Second, over-expression of Pygo2 facilitated the expression of P-glycoprotein, which acts as a drug efflux pump, by promoting the transcription of Multi-drug resistance 1 (MDR1) at the MDR1 promoter loci, resulting in acceleration of the efflux of PTX.

Protein arginine methyltransferase 1 may be involved in pregnane x receptor-activated overexpression of multidrug resistance 1 gene during acquired multidrug resistant.

PurposePregnane x receptor (PXR) - activated overexpression of the multidrug resistance 1 (MDR1) gene is an important way for tumor cells to acquire drug resistance. However, the detailed mechanism still remains unclear. In the present study, we aimed to investigate whether protein arginine methyl transferase 1(PRMT1) is involved in PXR - activated overexpression of MDR1 during acquired multidrug resistant.Experimental DesignArginine methyltransferase inhibitor 1 (AMI-1) was used to pharmacologically block PRMT1 in resistant breast cancer cells (MCF7/adr). The mRNA and protein levels of MDR1 were detected by real-time PCR and western blotting analysis. Immunofluorescence microscopy and co-immunoprecipitation were used to investigate the physical interaction between PXR and PRMT1. Then, 136 candidate compounds were screened for PRMT1 inhibitors. Lastly, luciferase reporter gene and nude mice bearing resistant breast cancer xenografts were adopted to investigate the anti-tumor effect of PRMT1 inhibitors when combined with adriamycin.ResultsAMI-1 significantly suppressed the expression of MDR1 in MCF7/adr cells and increased cells sensitivity of MCF7/adr to adriamycin. Physical interaction between PRMT1 and PXR exists in MCF7/adr cells, which could be disrupted by AMI-1. Those results suggest that PRMT1 may be involved in PXR-activated overexpression of MDR1 in resistant breast cancer cells, and AMI-1 may suppress MDR1 by disrupting the interaction between PRMT1 and PXR. Then, five compounds including rutin, isoquercitrin, salvianolic acid A, naproxen, and felodipline were identified to be PRMT1 inhibitors. Finally, those PRMT1 inhibitors were observed to significantly decrease MDR1 promoter activity in vitro and enhance the antitumor effect of adriamycin in nude mice that bearing resistant breast cancer xenografts.ConclusionsPRMT1 may be an important co-activator of PXR in activating MDR1 gene during acquired resistance, and PRMT1 inhibitor combined with chemotherapy drugs may be a new strategy for overcoming tumor MDR.

Sinapine reverses multi-drug resistance in MCF-7/dox cancer cells by downregulating FGFR4/FRS2α-ERK1/2 pathway-mediated NF-κB activation

BackgroundSinapine, an alkaloid derived from seeds of the cruciferous species, shows favorable biological properties, such as antioxidant and radio-protective activities. The inhibitory effect of sinapine on acquired chemoresistance in tumor cells and the underlying molecular mechanisms remain unknown. AimWe examined the effect of sinapine on reversal of chemoresistance in Michigan Cancer Foundation 7 (MCF-7)/dox breast cancer cells. ResultsCombination treatment with sinapine and doxorubicin synergistically increased the cytotoxicity of doxorubicin in MCF-7/dox cells, as shown using a cell apoptosis assay. An accumulation assay demonstrated that sinapine increased the intracellular concentration of doxorubicin in a dose-dependent manner. Immunoblotting and real time polymerase chain reaction (RT-PCR) analysis showed that sinapine downregulated multi-drug resistance 1 (MDR1) expression. A significant correlation was observed between the expression of MDR1, phospho-factor receptor substrate (FRS), phospho-extracellular signal regulated kinase (ERK)1/2, and nuclear factor kappa B (NF-κB). Chromatin immunoprecipitation (ChIP) assay indicated that sinapine inhibited binding of the transcription factor NF-κB to the MDR1 promoter. ConclusionsOur findings indicated that sinapine played an important role in the downregulation of MDR1 expression through suppression of fibroblast growth factor receptor (FGFR)4/FRS2α-ERK1/2 mediated NF-κB activation in MCF-7/dox cancer cells.

Etoposide enhances antitumor efficacy of MDR1-driven oncolytic adenovirus through autoupregulation of the MDR1 promoter activity.

Conditionally replicating adenoviruses (CRAds), or oncolytic adenoviruses, such as E1B55K-deleted adenovirus, are attractive anticancer agents. However, the therapeutic efficacy of E1B55K-deleted adenovirus for refractory solid tumors has been limited. Environmental stress conditions may induce nuclear accumulation of YB-1, which occurs in multidrug-resistant and adenovirus-infected cancer cells. Overexpression and nuclear localization of YB-1 are associated with poor prognosis and tumor recurrence in various cancers. Nuclear YB-1 transactivates the multidrug resistance 1 (MDR1) genes through the Y-box. Here, we developed a novel E1B55K-deleted adenovirus driven by the MDR1 promoter, designed Ad5GS3. We tested the feasibility of using YB-1 to transcriptionally regulate Ad5GS3 replication in cancer cells and thereby to enhance antitumor efficacy. We evaluated synergistic antitumor effects of oncolytic virotherapy in combination with chemotherapy. Our results show that adenovirus E1A induced E2F-1 activity to augment YB-1 expression, which shut down host protein synthesis in cancer cells during adenovirus replication. In cancer cells infected with Ad5WS1, an E1B55K-deleted adenovirus driven by the E1 promoter, E1A enhanced YB-1 expression, and then further phosphorylated Akt, which, in turn, triggered nuclear translocation of YB-1. Ad5GS3 in combination with chemotherapeutic agents facilitated nuclear localization of YB-1 and, in turn, upregulated the MDR1 promoter activity and enhanced Ad5GS3 replication in cancer cells. Thus, E1A, YB-1, and the MDR1 promoter form a positive feedback loop to promote Ad5GS3 replication in cancer cells, and this regulation can be further augmented when chemotherapeutic agents are added. In the in vivo study, Ad5GS3 in combination with etoposide synergistically suppressed tumor growth and prolonged survival in NOD/SCID mice bearing human lung tumor xenografts. More importantly, Ad5GS3 exerted potent oncolytic activity against clinical advanced lung adenocarcinoma, which was associated with elevated levels of nuclear YB-1 and cytoplasmic MDR1 expression in the advanced tumors. Therefore, Ad5GS3 may have therapeutic potential for cancer treatment, especially in combination with chemotherapy. Because YB-1 is expressed in a broad spectrum of cancers, this oncolytic adenovirus may be broadly applicable.

ZHX2 enhances the cytotoxicity of chemotherapeutic drugs in liver tumor cells by repressing MDR1 via interfering with NF-YA.

We previously reported the tumor suppressor function of Zinc-fingers and homeoboxes 2 (ZHX2) in hepatocellular carcinoma (HCC). Other studies indicate the association of increased ZHX2 expression with improved response to high dose chemotherapy in multiple myeloma. Here, we aim to test whether increased ZHX2 levels in HCC cells repress multidrug resistance 1(MDR1) expression resulting in increased sensitivity to chemotherapeutic drugs. We showed evidence that increased ZHX2 levels correlated with reduced MDR1 expression and enhanced the cytotoxicity of CDDP and ADM in different HCC cell lines. Consistently, elevated ZHX2 significantly reduced ADM efflux in HepG2 cells and greatly increased the CDDP-mediated suppression of liver tumor growth in vivo. Furthermore, immunohistochemical staining demonstrated the inverse correlation of ZHX2 and MDR1 expression in HCC tissues. Luciferase report assay showed that ZHX2 repressed the MDR1 promoter activity, while knockdown of NF-YA or mutating the NF-Y binding site eliminated this ZHX2-mediated repression of MDR1 transcription. Co-IP and ChIP assay further suggested that ZHX2 interacted with NF-YA and reduced NF-Y binding to the MDR1 promoter. Taken together, we clarify that ZHX2 represses NF-Y-mediated activation of MDR1 transcription and, in doing so, enhances the effects of chemotherapeutics in HCC cells both in vitro and in vivo.

Dioscin enhances methotrexate absorption by down-regulating MDR1 in vitro and in vivo

The purpose of this study was to investigate the enhancing effect of dioscin on the absorption of methotrexate (MTX) and clarify the molecular mechanism involved in vivo and in vitro. Dioscin increased MTX chemosensitivity and transepithelial flux in the absorptive direction, significantly inhibiting multidrug resistance 1 (MDR1) mRNA and protein expression and MDR1 promoter and nuclear factor κ-B (NF-κB) activities in Caco-2 cells. Moreover, inhibitor κB-α (IκB-α) degradation was inhibited by dioscin. Dioscin enhanced the intracellular concentration of MTX by down-regulating MDR1 expression through a mechanism that involves NF-κB signaling pathway inhibition in Caco-2 cells. Dioscin strengthened MTX absorption by inhibiting MDR1 expression in rat intestine. In addition, even though MTX is absorbed into the enterocytes, there was no increase in toxicity observed, and that, in fact, decreased toxicity was seen.

Expression of Aryl Hydrocarbon Receptor Nuclear Translocator Enhances Cisplatin Resistance by Upregulating MDR1 Expression in Cancer Cells

The identification of molecular pathways in cancer cells is important for understanding the cells' underlying biology and for designing effective cancer therapies. We demonstrate that the expression of aryl hydrocarbon receptor nuclear translocator (ARNT) is critical during the development of cisplatin resistance. The reduced expression of ARNT was correlated with cisplatin-induced cell death in drug-sensitive cells. In addition, suppression of ARNT reversed the characteristics of cisplatin-resistant cells, making these cells cisplatin-sensitive, and significantly enhanced caspase-3 activation, DNA fragmentation, and apoptosis. The inhibition of colony formation, regulated by cisplatin, was more significant in ARNT-knockdown cells than in parental cells. In a xenograft analysis of severe combined immunodeficiency mice, cisplatin also efficiently inhibited ARNT-deficient c4 tumors but not ARNT-containing vT2 tumor formation. Furthermore, the downregulation of multidrug resistance 1 (MDR1) expression and retention of drugs in cells caused by suppression of ARNT, resulting in the resensitization of drug-resistant cells to cisplatin, was observed. When overexpressed, ARNT interacted with Sp1 to enhance the expression of MDR1 through Sp1-binding sites on the MDR1 promoter, resulting in a reversal of the effect of cisplatin on cell death. In addition, ARNT-induced MDR1 expression was inhibited in Sp1-knockdown cells. These results reveal previously unrecognized, multifaceted functions of ARNT in establishing the drug-resistant properties of cancer cells by the upregulation of MDR1, highlighting ARNT's potential as a therapeutic target in an important subset of cancers.

Dioscin Restores the Activity of the Anticancer Agent Adriamycin in Multidrug-Resistant Human Leukemia K562/Adriamycin Cells by Down-Regulating MDR1 via a Mechanism Involving NF-κB Signaling Inhibition

The purpose of this study was to investigate the ameliorating effect of dioscin (1) on multidrug resistance (MDR) in adriamycin (ADR)-resistant erythroleukemic cells (K562/adriamycin, K562/ADR) and to clarify the molecular mechanisms involved. High levels of multidrug resistance 1 (MDR1) mRNA and protein and reduced ADR retention were found in K562/ADR cells compared with parental cells (K562). Dioscin (1), a constituent of plants in the genus Discorea, significantly inhibited MDR1 mRNA and protein expression and MDR1 promoter and nuclear factor κ-B (NF-κB) activity in K562/ADR cells. MDR1 mRNA and protein suppression resulted in the subsequent recovery of intracellular drug accumulation. Additionally, inhibitor κB-α (IκB-α) degradation was inhibited by 1. Dioscin (1) reversed ADR-induced MDR by down-regulating MDR1 expression by a mechanism that involves the inhibition of the NF-κB signaling pathway. These findings provide evidence to support the further investigation of the clinical application of dioscin (1) as a chemotherapy adjuvant.

Egr-1 enhances drug resistance of breast cancer by modulating MDR1 expression in a GGPPS-independent manner

The multidrug resistance 1 (MDR1) gene product P-glycoprotein is an ATP-dependent efflux pump associated with chemotherapy failure in breast cancer. In the present study, we show that paclitaxel induces MDR1 expression in the MCF-7 breast cancer cell line in a MAPK/Egr-1-dependent manner. Paclitaxel exposure activated the Erk1/2/MAPK pathway and promoted the accumulation of the early response transcription factor Egr-1 in MCF-7 cells. Egr-1 binds to the GC element on the proximal MDR1 promoter to enhance MDR1 transcription. Loss of Egr-1 function in paclitaxel-resistant MCF-7 cells decreased MDR1 expression, whereas inhibiting Erk1/2 activity reduced both Egr-1 accumulation and MDR1 expression. These findings suggest that Erk1/2-induced Egr-1 accumulation activates MDR1 transcription and thereby induces the drug resistance observed in paclitaxel-resistant MCF-7 cells. Further mechanistic studies indicate that Egr-1 most likely does not induce the constitutive activation of Erk1/2 through its target gene geranylgeranyl diphosphate synthase (GGPPS), which regulates Ras prenylation. Indeed, our results suggest a novel pathway by which paclitaxel induces MDR1 expression, possibly illuminating a potential target pathway for the prevention of MDR1-mediated drug resistance.

Clitocine Reversal of P-Glycoprotein Associated Multi-Drug Resistance through Down-Regulation of Transcription Factor NF-κB in R-HepG2 Cell Line

Multidrug resistance(MDR)is one of the major reasons for failure in cancer chemotherapy and its suppression may increase the efficacy of therapy. The human multidrug resistance 1 (MDR1) gene encodes the plasma membrane P-glycoprotein (P-gp) that pumps various anti-cancer agents out of the cancer cell. R-HepG2 and MES-SA/Dx5 cells are doxorubicin induced P-gp over-expressed MDR sublines of human hepatocellular carcinoma HepG2 cells and human uterine carcinoma MES-SA cells respectively. Herein, we observed that clitocine, a natural compound extracted from Leucopaxillus giganteus, presented similar cytotoxicity in multidrug resistant cell lines compared with their parental cell lines and significantly suppressed the expression of P-gp in R-HepG2 and MES-SA/Dx5 cells. Further study showed that the clitocine increased the sensitivity and intracellular accumulation of doxorubicin in R-HepG2 cells accompanying down-regulated MDR1 mRNA level and promoter activity, indicating the reversal effect of MDR by clitocine. A 5′-serial truncation analysis of the MDR1 promoter defined a region from position −450 to −193 to be critical for clitocine suppression of MDR1. Mutation of a consensus NF-κB binding site in the defined region and overexpression of NF-κB p65 could offset the suppression effect of clitocine on MDR1 promoter. By immunohistochemistry, clitocine was confirmed to suppress the protein levels of both P-gp and NF-κB p65 in R-HepG2 cells and tumors. Clitocine also inhibited the expression of NF-κB p65 in MES-SA/Dx5. More importantly, clitocine could suppress the NF-κB activation even in presence of doxorubicin. Taken together; our results suggested that clitocine could reverse P-gp associated MDR via down-regulation of NF-κB.

Metformin inhibits P‐glycoprotein expression via the NF‐κB pathway and CRE transcriptional activity through AMPK activation

The expression of P-glycoprotein (P-gp), encoded by the multidrug resistance 1 (MDR1) gene, is associated with the emergence of the MDR phenotype in cancer cells. We investigated whether metformin (1,1-dimethylbiguanide hydrochloride) down-regulates MDR1 expression in MCF-7/adriamycin (MCF-7/adr) cells. MCF-7 and MCF-7/adr cells were incubated with metformin and changes in P-gp expression were determined at the mRNA, protein and functional level. Transient transfection assays were performed to assess its gene promoter activities, and immunoblot analysis to study its molecular mechanisms of action. Metformin significantly inhibited MDR1 expression by blocking MDR1 gene transcription. Metformin also significantly increased the intracellular accumulation of the fluorescent P-gp substrate rhodamine-123. Nuclear factor-κB (NF-κB) activity and the level of IκB degradation were reduced by metformin treatment. Moreover, transduction of MCF-7/adr cells with the p65 subunit of NF-κB induced MDR1 promoter activity and expression, and this effect was attenuated by metformin. The suppression of MDR1 promoter activity and protein expression was mediated through metformin-induced activation of AMP-activated protein kinase (AMPK). Small interfering RNA methods confirmed that reduction of AMPK levels attenuates the inhibition of MDR1 activation associated with metformin exposure. Furthermore, the inhibitory effects of metformin on MDR1 expression and cAMP-responsive element binding protein (CREB) phosphorylation were reversed by overexpression of a dominant-negative mutant of AMPK. These results suggest that metformin activates AMPK and suppresses MDR1 expression in MCF-7/adr cells by inhibiting the activation of NF-κB and CREB. This study reveals a novel function of metformin as an anticancer agent.

Involvement of MyoD and PEA3 in regulation of transcription activity of MDR1 gene

Overexpression of multidrug resistance 1 (MDR1) in cancer remains one of the major causes for the failure of chemotherapy. In the present study, we found that MyoD and PEA3 could activate P-glycoprotein (P-gp) expression in SGC7901 cells. Knockdown of MyoD and PEA3 attenuated MDR1 expression and increased the sensitivity of multidrug resistant cancer cells to cytotoxic drugs that were transported by P-gp in SGC7901/VCR cells. MyoD or PEA3 could bind to the E-box and PEA3 sites on the MDR1 promoter and activate its transcription. The regulation of MDR1 expression by MyoD and PEA3 may provide potential ways to overcome MDR in cancer treatment.

Promoter Methylation Status of Multidrug Resistance 1 (MDR1) Gene in Noncancerous Gastric Mucosa Correlates With Helicobacter Pylori Infection and Gastric Cancer Occurrence

ABSTRACTWe investigated the methylation status of multidrug resistance 1 (MDR1) gene promoter in noncancerous gastric mucosa in relation to gastric cancer (GC) occurrence. Methylation of the MDR1 promoter was estimated in 127 GC and 82 non-GC patients. A significant association was found between higher methylation ratio and occurrence of GC. Higher methylation ratio was also associated with occurrence of GC in both Helicobacter pylori–positive and Helicobacter pylori–negative subjects. Higher MR was also associated with Helicobacter pylori infection. Methylation status of MDR1 gene in noncancerous gastric mucosa would be useful for predicting GC occurrence.

CpG hypomethylation of MDR1 gene in tumor and serum of invasive ductal breast carcinoma patients

Objectives: Multidrug resistance 1 ( MDR1) gene encodes P-glycoprotein (P-gp), a transmembrane calcium-dependent efflux pump, implicated in drug resistance. In this prospective study, methylation status of MDR1 promoter and its correlation with clinicopathological parameters were evaluated in tumor and serum of breast cancer patients. Design and methods: Methylation-specific PCR was carried out to investigate the promoter methylation status of MDR1 in tumor and serum of 100 patients with invasive ductal carcinomas of breast (IDCs). The effect of promoter methylation on protein expression was evaluated by immunohistochemistry. Results: MDR1 was hypomethylated in 47% tumors and 44% paired sera of IDC patients and correlated significantly with increased tumor size and advanced tumor stage. Promoter hypomethylation of MDR1 in serum DNA showed 98% specificity and 50% sensitivity. Conclusions: Hypomethylation of MDR1 promoter in IDCs accounted for P-gp overexpression and aggressive biologic behavior in a subset of patients. Detection of these epigenetic changes in circulating DNA may not only enhance insight into the biological behavior of the primary tumor of an individual but may also provide valuable information regarding prognosis that can be readily monitored throughout the disease course.

Cell type dependent regulation of multidrug resistance-1 gene expression by AML1-ETO

The AML1-ETO fusion protein is generated from the 8;21 chromosome translocation that is commonly identified in acute myeloid leukemia. AML1-ETO is a DNA binding transcription factor and has been demonstrated to play a critical role in promoting leukemogenesis. Therefore, it is important to define the molecular mechanism of AML1-ETO in the regulation of gene expression. Here, we report that the effect of AML1-ETO on the promoter of multidrug resistance-1 (MDR1) gene, a known AML1-ETO target, is highly cell type specific. Besides observing repression of the MDR1 promoter in C33A and CV-1 cells as reported previously, AML1-ETO strongly activated the promoter in K562 and B210 cells. More importantly, this activation required both the AML1 and ETO portions of the fusion protein, but did not depend on the AML1 binding site in MDR1 promoter. Furthermore, results from promoter deletion analysis and chromatin immunoprecipitation assays suggested that this activation effect was likely through the influence of the general transcription machinery rather than promoter-specific factors. Based on these data, we propose that AML1-ETO may have opposing effects on gene expression depending on the various conditions of the cellular environment.

PO2-dependent Differential Regulation of Multidrug Resistance 1 Gene Expression by the c-Jun NH2-terminal Kinase Pathway

Hypoxia-induced multidrug resistance 1 (MDR1) gene expression is known to be mediated by c-Jun NH(2)-terminal kinase (JNK) activation. However, the molecular mechanisms underlying this action of JNK remain elusive. On the contrary, there has been increasing evidence for a negative correlation of JNK activity with MDR1 expression under normoxic conditions. Here, we present evidence that the JNK pathway represses MDR1 expression in normoxia and activates MDR1 expression in hypoxia. Our data show that JNK pathway-induced MDR1 repression in normoxia is mediated by increased c-Jun binding to activator protein 1 site, located in the MDR1 promoter, and requires the activity of histone deacetylase 5. In contrast, JNK pathway-induced MDR1 activation in hypoxia is independent of the activator protein 1 site. Rather, this action is dependent on increased hypoxia-inducible factor 1 (HIF1) binding to the hypoxia response element in the MDR1 promoter, which is promoted by the interaction of HIF1alpha with c-Jun in the nucleus and requires the activity of the p300/CBP (CREB-binding protein) coactivator.

Riboregulator H19 induction of MDR1-associated drug resistance in human hepatocellular carcinoma cells

Acquisition of drug resistance is one of the main obstacles encountered in cancer chemotherapy. Overexpression of multi-drug resistance 1 (MDR1) gene and its protein product P-glycoprotein, accompanied with a decrease in doxorubicin accumulation level, was observed in doxorubicin-resistant R-HepG2 cells, a subline derived by selection of human hepatocellular carcinoma HepG2 cells with doxorubicin. In addition, Northern-blot analysis revealed an eight fold upregulation of the imprinted H19 mRNA in R-HepG2 cells. H19 knockdown by transfection with antisense H19 oligonucleotides suppressed the MDR1/P-glycoprotein expression, increased the cellular doxorubicin accumulation level and sensitized doxorubicin toxicity in both HepG2 parent cells and R-HepG2 cells. Results from methylation-specific polymerase chain reaction analysis indicated that the MDR1 gene promoter was hypomethylated in R-HepG2 cells. Antisense H19 oligonucleotides transfection induced a marked increase in the percentage of MDR1 promoter methylation and decrease in MDR1 expression in R-HepG2 cells. Thus, the H19 gene is believed to induce P-glycoprotein expression and MDR1-associated drug resistance at least in liver cancer cells through regulation of MDR1 promoter methylation.