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1.
J Mol Endocrinol ; 46(3): 205-16, 2011 Jun.
Article in English | MEDLINE | ID: mdl-21321095

ABSTRACT

Recently, crosstalk between sphingolipid signaling pathways and steroid hormones has been illuminated as a possible therapeutic target. Sphingosine kinase (SK), the key enzyme metabolizing pro-apoptotic ceramide to pro-survival sphingosine-1-phosphate (S1P), is a promising therapeutic target for solid tumor cancers. In this study, we examined the ability of pharmacological inhibition of S1P formation to block estrogen signaling as a targeted breast cancer therapy. We found that the Sphk1/2 selective inhibitor (SK inhibitor (SKI))-II, blocked breast cancer viability, clonogenic survival and proliferation. Furthermore, SKI-II dose-dependently decreased estrogen-stimulated estrogen response element transcriptional activity and diminished mRNA levels of the estrogen receptor (ER)-regulated genes progesterone receptor and steroid derived factor-1. This inhibitor binds the ER directly in the antagonist ligand-binding domain. Taken together, our results suggest that SKIs have the ability to act as novel ER signaling inhibitors in breast carcinoma.


Subject(s)
Breast Neoplasms/metabolism , Isoenzymes/metabolism , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Signal Transduction/drug effects , Thiazoles/pharmacology , Blotting, Western , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Computer Simulation , Estrogen Receptor alpha , Humans , Isoenzymes/antagonists & inhibitors , Mass Spectrometry , Phosphotransferases (Alcohol Group Acceptor)/antagonists & inhibitors , Reverse Transcriptase Polymerase Chain Reaction
2.
Endocrinology ; 151(11): 5124-35, 2010 Nov.
Article in English | MEDLINE | ID: mdl-20861237

ABSTRACT

Alterations in sphingolipid metabolism have been shown to contribute to the development of endocrine resistance and breast cancer tumor survival. Sphingosine kinase (SK), in particular, is overexpressed in breast cancer and is a promising target for breast cancer drug development. In this study, we used the novel SK inhibitor ABC294640 as a tool to explore the relationship between SK and estrogen (E2) receptor (ER) signaling in breast cancer cells. Treatment with ABC294640 decreased E2-stimulated ERE-luciferase activity in both MCF-7 and ER-transfected HEK293 cells. Furthermore, the inhibitor reduced E2-mediated transcription of the ER-regulated genes progesterone receptor and SDF-1. Competitive receptor-binding assays revealed that ABC294640 binds in the antagonist ligand-binding domain of the ER, acting as a partial antagonist similar to tamoxifen. Finally, treatment with ABC294640 inhibited ER-positive breast cancer tumor formation in vivo. After 15 d of treatment with ABC294640, tumor volume was reduced by 68.4% (P < 0.05; n = 5) compared with control tumors, with no marked weight loss or illness. Taken together, these results provide strong evidence that this novel SK inhibitor, which had not previously been known to interact with E2 signaling pathways, has therapeutic potential in treating ER-positive breast cancer via inhibition of both SK and ER signaling.


Subject(s)
Adamantane/analogs & derivatives , Cell Proliferation/drug effects , Estradiol/metabolism , Phosphotransferases (Alcohol Group Acceptor)/antagonists & inhibitors , Pyridines/pharmacology , Receptors, Estrogen/metabolism , Adamantane/pharmacology , Analysis of Variance , Animals , Blotting, Western , Cell Line , Cell Survival/drug effects , Cells, Cultured , Female , Gene Expression/drug effects , Humans , Immunohistochemistry , Mice , Phosphorylation/drug effects , Phosphotransferases (Alcohol Group Acceptor)/genetics , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Receptors, Estrogen/genetics , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/drug effects , Transcription, Genetic/drug effects
3.
Exp Biol Med (Maywood) ; 234(11): 1253-63, 2009 Nov.
Article in English | MEDLINE | ID: mdl-19546354

ABSTRACT

An estimated 182,640 women and 1,990 men were diagnosed with breast cancer in 2008, and approximately 40,480 women and 450 men died from the disease. Thus, continued mechanistic studies are needed to understand the causes and develop additional therapeutics for this complicated disease. The MCF-7 cell system is one of the most recognized models for estrogen receptor (ER)-positive breast cancer and has generated approximately 13,000 publications cited in PubMed to date. A number of clues for biological mechanisms related to apoptotic/anti-apoptotic pathways and chemoresistance were elucidated and summarized in our previous review. The focus of this review is new knowledge of the central role of sphingolipid signaling in apoptotic mechanisms in estrogen receptor-positive breast cancer. The ultimate goal is to target crucial steps in survival signaling pathways that may ultimately provide additional translational solutions to the successful pharmacologic treatment of breast cancer.


Subject(s)
Apoptosis , Drug Resistance, Neoplasm , Models, Biological , Sphingolipids/metabolism , Breast Neoplasms/enzymology , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Breast Neoplasms/therapy , Cell Line, Tumor , Female , Humans , Tumor Necrosis Factor-alpha/metabolism
4.
Mol Ther ; 17(3): 430-8, 2009 Mar.
Article in English | MEDLINE | ID: mdl-19107118

ABSTRACT

Radiation resistance in a subset of prostate tumors remains a challenge to prostate cancer radiotherapy. The current study on the effects of radiation on prostate cancer cells reveals that radiation programs an unpredicted resistance mechanism by upregulating acid ceramidase (AC). Irradiated cells demonstrated limited changes of ceramide levels while elevating levels of sphingosine and sphingosine-1-phosphate. By genetically downregulating AC with small interfering RNA (siRNA), we observed radiosensitization of cells using clonogenic and cytotoxicity assays. Conversely, AC overexpression further decreased sensitivity to radiation. We also observed that radiation-induced AC upregulation was sufficient to create cross-resistance to chemotherapy as demonstrated by decreased sensitivity to Taxol and C(6) ceramide compared to controls. Lower levels of caspase 3/7 activity were detected in cells pretreated with radiation, also indicating increased resistance. Finally, utilization of the small molecule AC inhibitor, LCL385, sensitized PPC-1 cells to radiation and significantly decreased tumor xenograft growth. These data suggest a new mechanism of cancer cell resistance to radiation, through upregulation of AC that is, in part, mediated by application of the therapy itself. An improved understanding of radiotherapy and the application of combination therapy achieved in this study offer new opportunities for the modulation of radiation effects in the treatment of cancer.


Subject(s)
Acid Ceramidase/metabolism , Prostatic Neoplasms/enzymology , Radiation-Sensitizing Agents/pharmacology , Up-Regulation/drug effects , Up-Regulation/radiation effects , Acid Ceramidase/antagonists & inhibitors , Acid Ceramidase/genetics , Animals , Cell Line, Tumor , Ceramides/metabolism , Enzyme Activation/drug effects , Enzyme Inhibitors/pharmacology , Humans , Male , Mice , Mice, Nude , Myristates/pharmacology , Paclitaxel/pharmacology , Phosphotransferases (Alcohol Group Acceptor)/metabolism , Propanolamines/pharmacology , Prostatic Neoplasms/genetics , RNA, Small Interfering/genetics , Sensitivity and Specificity , Sphingosine/analogs & derivatives , Sphingosine/metabolism , Xenograft Model Antitumor Assays
5.
Cancer Biol Ther ; 6(9): 1455-60, 2007 Sep.
Article in English | MEDLINE | ID: mdl-17881906

ABSTRACT

Among the many processes regulating cell death, ceramide signaling is a vital component. We previously determined that acid ceramidase (AC) is upregulated in 60% of primary prostate cancer (PCa) tissues, suggesting that AC may play a role in tumor development. In order to determine the significance of AC elevation, stable clones of DU145 cells with AC overexpression (AC-EGFP) were generated. Compared to controls (EGFP), AC-EGFP cells exhibited enhanced cell proliferation and migration. Subcutaneous injection of AC-EGFP cells into Nu/Nu mice resulted in larger tumor volumes compared to EGFP controls. Moreover, using the MTS viability assay, AC-EGFP cells were more resistant to cell death induced by doxorubicin, cisplatin, etoposide, gemcitabine or C6-ceramide. Conversely, knock down of AC using siRNA, sensitized AC-EGFP cells to these drugs. In addition, mass spectroscopic analysis of sphingolipids indicated that long chain ceramide levels were decreased in AC-EGFP cells treated with either doxorubicin or etoposide. In conclusion, this study implicates AC as a critical regulator of PCa progression by affecting not only tumor cell proliferation and migration but also responses to drug therapy, suggesting AC as a potential therapeutic target in advanced PCa.


Subject(s)
Drug Resistance, Neoplasm , Galactosylgalactosylglucosylceramidase/biosynthesis , Gene Expression Regulation, Neoplastic , Prostatic Neoplasms/drug therapy , Prostatic Neoplasms/enzymology , Animals , Cell Line, Tumor , Cell Movement , Cell Proliferation , Cell Survival , Ceramides/metabolism , Disease Progression , Humans , Male , Mice , Mice, Nude
6.
Mol Ther ; 15(7): 1259-63, 2007 Jul.
Article in English | MEDLINE | ID: mdl-17426710

ABSTRACT

Head and neck squamous cell cancers (HNSCC) are particularly aggressive and are resistant to many forms of treatment. Ceramide metabolism has been shown to play an important role in cancer progression and cancer resistance to therapy in many tumor models, including HNSCC. Here, we study the role of the ceramide-metabolizing enzyme acid ceramidase (AC) in therapeutic responses in HNSCC. First, we show that AC is over-expressed in 70% of head and neck squamous cell tumors compared with normal tissues, suggesting that this enzyme may play an important role in facilitating HNSCC growth. Next, comparison of three HNSCC cell lines with low, medium, and high levels of AC reveals an inverse correlation between the levels of AC and their response to exogenous C-6-ceramide. Furthermore, over-expression of AC in SCC-1 cells increased resistance to Fas-induced cell killing. Conversely, down-regulation of AC using specific AC small interfering RNA (siRNA) sensitized the SCC-1 cancer cell line to Fas-induced apoptosis. Finally, we show that the AC inhibitor LCL 204 can sensitize HNSCC cell lines to Fas-induced apoptosis both in vitro and in a xenograft model in vivo, suggesting that the combination of FasL gene therapy and LCL 204 may become a new treatment option for advanced-stage head and neck cancer.


Subject(s)
Enzyme Inhibitors/pharmacology , Fas Ligand Protein/genetics , Fas Ligand Protein/metabolism , Galactosylgalactosylglucosylceramidase/antagonists & inhibitors , Galactosylgalactosylglucosylceramidase/metabolism , Genetic Therapy , Head and Neck Neoplasms/metabolism , Animals , Cell Line, Tumor , Cell Survival , Ceramides/metabolism , Down-Regulation , Female , Gene Expression Regulation, Neoplastic , Head and Neck Neoplasms/genetics , Humans , Mice , Mice, Nude , RNA, Small Interfering/genetics , Sensitivity and Specificity , Xenograft Model Antitumor Assays
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