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Analysis Of The Regulation And Function Of Cip2a To Identify Candidate Biomarkers For Prostate Cancer, Diana Savoly

Graduate School of Biomedical Sciences Theses and Dissertations

Protein Phosphatase 2A (PP2A) is a tumor suppressor involved in the regulation of several signaling pathways and the cell cycle. PP2A becomes inactivated by several inhibitors, including Cancerous Inhibitor of PP2A (CIP2A). CIP2A has been identified as an oncogene, which is over-expressed in cancers and inhibits PP2A through direct interaction. CIP2A is recognized as a biomarker for cancer; however, it is not cancer-specific. Therefore, we identified and examined the use of CIP2A-regulated proteins as potential biomarkers in prostate cancer to better diagnose prostate cancer in patients. Currently, Prostate Specific Antigen (PSA) is widely used to detect prostate cancer; however, it …

The Tumor Suppressor Tere1 (Ubiad1) Prenyltransferase Regulates The Elevated Cholesterol Phenotype In Castration Resistant Prostate Cancer By Controlling A Program Of Ligand Dependent Sxr Target Genes., William J. Fredericks, Jorge Sepulveda, Priti Lai, John E. Tomaszewski, Ming-Fong Lin, Terry Mcgarvey, Frank J. Rauscher, S. Bruce Malkowicz

Journal Articles: Biochemistry & Molecular Biology

Castrate-Resistant Prostate Cancer (CRPC) is characterized by persistent androgen receptor-driven tumor growth in the apparent absence of systemic androgens. Current evidence suggests that CRPC cells can produce their own androgens from endogenous sterol precursors that act in an intracrine manner to stimulate tumor growth. The mechanisms by which CRPC cells become steroidogenic during tumor progression are not well defined. Herein we describe a novel link between the elevated cholesterol phenotype of CRPC and the TERE1 tumor suppressor protein, a prenyltransferase that synthesizes vitamin K-2, which is a potent endogenous ligand for the SXR nuclear hormone receptor. We show that 50% …

Impaired Expression Of Protein Phosphatase 2a Subunits Enhances Metastatic Potential Of Human Prostate Cancer Cells Through Activation Of Akt Pathway., P Pandey, Parthasarathy Seshacharyulu, Srustidhar Das, Satyanarayana Rachagani, Moorthy P. Ponnusamy, Y Yan, Sonny L. Johansson, K Datta, Ming-Fong Lin, Surinder K. Batra

Journal Articles: Biochemistry & Molecular Biology

BACKGROUND: Protein phosphatase 2A (PP2A) is a dephosphorylating enzyme, loss of which can contribute to prostate cancer (PCa) pathogenesis. The aim of this study was to analyse the transcriptional and translational expression patterns of individual subunits of the PP2A holoenzyme during PCa progression.

METHODS: Immunohistochemistry (IHC), western blot, and real-time PCR was performed on androgen-dependent (AD) and androgen-independent (AI) PCa cells, and benign and malignant prostate tissues for all the three PP2A (scaffold, regulatory, and catalytic) subunits. Mechanistic and functional studies were performed using various biochemical and cellular techniques.

RESULTS: Through immunohistochemical analysis we observed significantly reduced levels of PP2A-A …

Androgens Upregulate Cdc25c Protein By Inhibiting Its Proteasomal And Lysosomal Degradation Pathways., Yu-Wei Chou, Li Zhang, Sakthivel Muniyan, Humera Ahmad, Satyendra Kumar, Syed Mahfuzul Alam, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

Cdc25C is a cell cycle protein of the dual specificity phosphatase family essential for activating the cdk1/Cyclin B1 complex in cells entering into mitosis. Since altered cell cycle is a hallmark of human cancers, we investigated androgen regulation of Cdc25C protein in human prostate cancer (PCa) cells, including androgen-sensitive (AS) LNCaP C-33 cells and androgen-independent (AI) LNCaP C-81 as well as PC-3 cells. In the regular culture condition containing fetal bovine serum (FBS), Cdc25C protein levels were similar in these PCa cells. In a steroid-reduced condition, Cdc25C protein was greatly decreased in AS C-33 cells but not AI C-81 or …

Marked Improvement Of Cytotoxic Effects Induced By Docetaxel On Highly Metastatic And Androgen-Independent Prostate Cancer Cells By Downregulating Macrophage Inhibitory Cytokine-1., M Mimeault, Sonny L. Johansson, Surinder K. Batra

Journal Articles: Biochemistry & Molecular Biology

BACKGROUND: Overexpression of macrophage inhibitory cytokine-1 (MIC-1) frequently occurs during the progression of prostate cancer (PC) to androgen-independent (AI) and metastatic disease states and is associated with a poor outcome of patients.

METHODS: The gain- and loss-of-function analyses of MIC-1 were performed to establish its implications for aggressive and chemoresistant phenotypes of metastatic and AI PC cells and the benefit of its downregulation for reversing docetaxel resistance.

RESULTS: The results have indicated that an enhanced level of secreted MIC-1 protein in PC3 cells is associated with their acquisition of epithelial-mesenchymal transition features and higher invasive capacity and docetaxel resistance. Importantly, …

Retinoids Regulate The Formation And Degradation Of Gap Junctions In Androgen-Responsive Human Prostate Cancer Cells., Linda Kelsey, Parul Katoch, Kristen E. Johnson, Surinder K. Batra, Parmender P. Mehta

Journal Articles: Biochemistry & Molecular Biology

The retinoids, the natural or synthetic derivatives of Vitamin A (retinol), are essential for the normal development of prostate and have been shown to modulate prostate cancer progression in vivo as well as to modulate growth of several prostate cancer cell lines. 9-cis-retinoic acid and all-trans-retinoic acid are the two most important metabolites of retinol. Gap junctions, formed of proteins called connexins, are ensembles of intercellular channels that permit the exchange of small growth regulatory molecules between adjoining cells. Gap junctional communication is instrumental in the control of cell growth. We examined the effect of 9-cis-retinoic acid and all-trans retinoic …

Pathobiological Implications Of The Expression Of Egfr, Pakt, Nf-Κb And Mic-1 In Prostate Cancer Stem Cells And Their Progenies., Murielle Mimeault, Sonny L. Johansson, Surinder K. Batra

Journal Articles: Biochemistry & Molecular Biology

The progression of prostate cancers (PCs) to locally invasive, androgen-independent and metastatic disease states is generally associated with treatment resistance and disease relapse. The present study was undertaken to establish the possibility of using a combination of specific oncogenic products, including epidermal growth factor receptor (EGFR), pAkt, nuclear factor-kappaB (NF-κB) and macrophage inhibitory cytokine-1 (MIC-1) as biomarkers and therapeutic targets for optimizing the management of patients with localized PC at earlier disease stages. The immunohistochemical and immunofluorescence data have revealed that the expression levels of EGFR, Ser(473)-pAkt, NF-κB p65 and MIC-1 proteins were significantly enhanced in the same subset of …

Histone Deacetylase Inhibitor Valproic Acid Suppresses The Growth And Increases The Androgen Responsiveness Of Prostate Cancer Cells., Yu-Wei Chou, Nagendra K. Chaturvedi, Shougiang Ouyang, Fen-Fen Lin, Dharam Kaushik, Jue Wang, Isaac Kim, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

We identified the molecular target by histone deacetylase (HDAC) inhibitors for exploring their potential prostate cancer (PCa) therapy. Upon HDAC inhibitors-treatment, LNCaP cell growth was suppressed, correlating with increased cellular prostatic acid phosphatase (cPAcP) expression, an authentic protein tyrosine phosphatase. In those cells, ErbB-2 was dephosphorylated, histone H3/H4 acetylation and methylation increased and cyclin proteins decreased. In PAcP shRNA-transfected C-81 cells, valproic acid (VPA) efficacy of growth suppression was diminished. Further, VPA pre-treatment enhanced androgen responsiveness of C-81, C4-2 and MDA PCa2b-AI cells. Thus, cPAcP expression is involved in growth suppression by HDAC inhibitors in PCa cells, and VPA pre-treatments …

The Raf/Mek/Extracellular Signal-Regulated Kinase 1/2 Pathway Can Mediate Growth Inhibitory And Differentiation Signaling Via Androgen Receptor Downregulation In Prostate Cancer Cells., Seung-Keun Hong, Jin-Hwan Kim, Ming-Fong Lin, Jong-In Park

Journal Articles: Biochemistry & Molecular Biology

Upregulated ERK1/2 activity is correlated with androgen receptor (AR) downregulation in certain prostate cancer (PCa) that exhibits androgen deprivation-induced neuroendocrine differentiation, but its functional relevance requires elucidation. We found that sustained ERK1/2 activation using active Raf or MEK1/2 mutants is sufficient to induce AR downregulation at mRNA and protein levels in LNCaP. Downregulation of AR protein, but not mRNA, was blocked by proteasome inhibitors, MG132 and bortezomib, indicating that the pathway regulation is mediated at multiple points. Ectopic expression of a constitutively active AR inhibited Raf/MEK/ERK-mediated regulation of the differentiation markers, neuron-specific enolase and neutral endopeptidase, and the cyclin-dependent kinase …

Tnfα Enhances The Motility And Invasiveness Of Prostatic Cancer Cells By Stimulating The Expression Of Selective Glycosyl- And Sulfotransferase Genes Involved In The Synthesis Of Selectin Ligands., Prakash Radhakrishnan, Vishwanath Chachadi, Ming-Fong Lin, Rakesh Singh, Reiji Kannagi, Pi-Wan Cheng

Journal Articles: Biochemistry & Molecular Biology

Sialyl Lewis x (sLe(x)) plays an important role in cancer metastasis. But, the mechanism for its production in metastatic cancers remains unclear. The objective of current study was to examine the effects of a proinflammatory cytokine on the expression of glycosyltransferase and sulfotransferase genes involved in the synthesis of selectin ligands in a prostate cancer cell line. Androgen-independent human lymph node-derived metastatic prostate cancer cells (C-81 LNCaP), which express functional androgen receptor and mimic the castration-resistant advanced prostate cancer, were used. TNFα treatment of these cells increased their binding to P-, E- and L-selectins, anti-sLe(x) antibody, and anti-6-sulfo-sialyl Lewis x …

Steroids Up-Regulate P66shc Longevity Protein In Growth Regulation By Inhibiting Its Ubiquitination., Santosh Kumar, Satyendra Kumar, Mythilypriya Rajendran, Syed Mahfuzul Alam, Fen-Fen Lin, Pi-Wan Cheng, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

BACKGROUND: p66Shc, an isoform of Shc adaptor proteins, mediates diverse signals, including cellular stress and mouse longevity. p66Shc protein level is elevated in several carcinomas and steroid-treated human cancer cells. Several lines of evidence indicate that p66Shc plays a critical role in steroid-related carcinogenesis, and steroids play a role in its elevated levels in those cells without known mechanism.

METHODS AND FINDINGS: In this study, we investigated the molecular mechanism by which steroid hormones up-regulate p66Shc protein level. In steroid-treated human prostate and ovarian cancer cells, p66Shc protein levels were elevated, correlating with increased cell proliferation. These steroid effects on …

P66shc--A Longevity Redox Protein In Human Prostate Cancer Progression And Metastasis : P66shc In Cancer Progression And Metastasis., Mythilypriya Rajendran, Paul Thomes, Li Zhang, Suresh Veeramani, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

p66Shc, a 66 kDa proto-oncogene Src homologous-collagen homologue (Shc) adaptor protein, is classically known in mediating receptor tyrosine kinase signaling and recently identified as a sensor to oxidative stress-induced apoptosis and as a longevity protein in mammals. The expression of p66Shc is decreased in mice and increased in human fibroblasts upon aging and in aging-related diseases, including prostate cancer. p66Shc protein level correlates with the proliferation of several carcinoma cells and can be regulated by steroid hormones. Recent advances point that p66Shc protein plays a role in mediating cross-talk between steroid hormones and redox signals by serving as a common …

Suppression Of Erbb-2 In Androgen-Independent Human Prostate Cancer Cells Enhances Cytotoxic Effect By Gemcitabine In An Androgen-Reduced Environment., Li Zhang, Jeffrey S. Davis, Stanislav Zelivianski, Fen-Fen Lin, Rachel Schutte, Thomas L. Davis, Ralph Hauke, Surinder K. Batra, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

We examined the efficacy of combination treatments utilizing cytotoxic drugs plus inhibitors to members of the ErbB-ERK signal pathway in human prostate cancer (PCa) LNCaP C-81 cells. Under an androgen-reduced condition, 50nM gemcitabine caused about 40% growth suppression on C-81 cells. Simultaneous treatment of gemcitabine plus 10microM AG825 produced 60% suppression (p

Upregulation Of Pip3-Dependent Rac Exchanger 1 (P-Rex1) Promotes Prostate Cancer Metastasis., Jianbing Qin, Yan Xie, Bo Wang, Mikio Hoshino, Dennis W. Wolff, Jing Zhao, Margaret A. Scofield, Frank J. Dowd, Ming-Fong Lin, Yaping Tu

Journal Articles: Biochemistry & Molecular Biology

Excessive activation of G-protein-coupled receptor (GPCR) and receptor tyrosine kinase (RTK) pathways has been linked to prostate cancer metastasis. Rac activation by guanine nucleotide exchange factors (GEFs) plays an important role in directional cell migration, a critical step of tumor metastasis cascades. We found that the upregulation of P-Rex1, a Rac-selective GEF synergistically activated by Gbetagamma freed during GPCR signaling, and PIP3, generated during either RTK or GPCR signaling, strongly correlates with metastatic phenotypes in both prostate cancer cell lines and human prostate cancer specimens. Silencing endogenous P-Rex1 in metastatic prostate cancer PC-3 cells selectively inhibited Rac activity and reduced …

Elevated Expression Of L-Selectin Ligand In Lymph Node-Derived Human Prostate Cancer Cells Correlates With Increased Tumorigenicity., Prakash Radhakrishnan, Ming-Fong Lin, Pi-Wan Cheng

Journal Articles: Biochemistry & Molecular Biology

Human prostate cancer LNCaP cells including C-33 and C-81 cells were originally derived from the lymph nodes of a patient with metastatic prostate cancer. These two cells were employed for characterization of L-selectin ligand and in vitro tumorigenicity, because they mimic the clinical conditions of early and late-stage human prostate cancer. C-81 cells exhibit higher in vitro migratory and invasive properties as compared with C-33 cells. We find that the L-selectin ligand and mucin glycan-associated MECA-79 epitope were elevated in C-81 cells. An increase of these glycotopes positively correlates with elevated tumorigenicity and expression of key glycosyl- and sulfotransferase genes. …

Androgen-Independent Prostate Cancer Cells Acquire The Complete Steroidogenic Potential Of Synthesizing Testosterone From Cholesterol., Paulette R. Dillard, Ming-Fong Lin, Shafiq A. Khan

Journal Articles: Biochemistry & Molecular Biology

The proliferation and differentiation of normal prostate epithelial cells depends upon the action of androgens produced by the testis. Prostate cancers retain the ability to respond to androgens in the initial stages of cancer development, but progressively become independent of exogenous androgens in advanced stages of the disease while maintaining the expression of functional androgen receptor (AR). In the present study, we have determined the potential of prostate cancer cells to synthesize androgens from cholesterol which may be involved in intracrine regulation of AR in advanced stages of the disease. Established androgen-independent prostate cancer cell lines, PC3 and DU145 cells, …

Mitochondrial Redox Signaling By P66shc Is Involved In Regulating Androgenic Growth Stimulation Of Human Prostate Cancer Cells., Suresh Veeramani, Ta-Chun Yuan, Fen-Fen Lin, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

p66Shc is shown to negatively regulate the life span in mice through reactive oxygen species (ROS) production. Recent reports, however, revealed that p66Shc protein level is significantly elevated in several human cancer tissues and growth-stimulated carcinoma cells, suggesting a mitogenic and carcinogenic role for p66Shc. In this communication, we demonstrate for the first time that p66Shc mediates androgenic growth signals in androgen-sensitive human prostate cancer cells through mitochondrial ROS production. Growth stimulation of prostate cancer cells with 5alpha-dihydrotestosterone (DHT) is accompanied by increased p66Shc level and ROS production, which is abolished by antioxidant treatments. However, antioxidant treatments do not affect …

Genome-Wide Expression Profiling Reveals Transcriptomic Variation And Perturbed Gene Networks In Androgen-Dependent And Androgen-Independent Prostate Cancer Cells., Ajay P. Singh, Sangeeta Bafna, Kunal Chaudhary, Ganesh Venkatraman, Lynette Smith, James D. Eudy, Sonny L. Johansson, Ming-Fong Lin, Surinder K. Batra

Journal Articles: Biochemistry & Molecular Biology

Previously, we have developed a unique in vitro LNCaP cell model, which includes androgen-dependent (LNCaP-C33), androgen-independent (LNCaP-C81) and an intermediate phenotype (LNCaP-C51) cell lines resembling the stages of prostate cancer progression to hormone independence. This model is advantageous in overcoming the heterogeneity associated with the prostate cancer up to a certain extent. We characterized and compared the gene expression profiles in LNCaP-C33 (androgen-dependent) and LNCaP-C81 (androgen-independent) cells using Affymetrix GeneChip array analyses. Multiple genes were identified exhibiting differential expression during androgen-independent progression. Among the important genes upregulated in androgen-independent cells were PCDH7, TPTE, TSPY, EPHA3, HGF, MET, EGF, TEM8, etc., …

Androgen-Regulated Formation And Degradation Of Gap Junctions In Androgen-Responsive Human Prostate Cancer Cells., Shalini Mitra, Lakshmanan Annamalai, Souvik Chakraborty, Kristen E. Johnson, Xiao-Hong Song, Surinder K. Batra, Parmender P. Mehta

Journal Articles: Biochemistry & Molecular Biology

The constituent proteins of gap junctions, called connexins (Cxs), have a short half-life. Despite this, the physiological stimuli that control the assembly of Cxs into gap junctions and their degradation have remained poorly understood. We show here that in androgen-responsive human prostate cancer cells, androgens control the expression level of Cx32-and hence the extent of gap junction formation-post-translationally. In the absence of androgens, a major fraction of Cx32 is degraded presumably by endoplasmic reticulum-associated degradation, whereas in their presence, this fraction is rescued from degradation. We also show that Cx32 and Cx43 degrade by a similar mechanism. Thus, androgens regulate …

Transcriptional Activation Of The Human Prostatic Acid Phosphatase Gene By Nf-Kappab Via A Novel Hexanucleotide-Binding Site., Stanislav Zelivianski, Richard Glowacki, Ming-Fong Lin

Journal Articles: Biochemistry & Molecular Biology

Human prostatic acid phosphatase (PAcP) is a prostate epithelium-specific differentiation antigen. Cellular PAcP functions as a neutral protein tyrosine phosphatase and is involved in regulating androgen-promoted prostate cancer cell proliferation. Despite the fact that the promoter of the PAcP gene has been cloned, the transcriptional factors that regulate PAcP expression remain unidentified. This article describes our analyses of the promoter of the PAcP gene. Deletion analyses of the promoter sequence up to -4893 (-4893/+87) revealed that a 577 bp fragment (-1356/-779) represents the unique positive cis-active element in human prostate cancer cells but not in HeLa cervix carcinoma cells. Interestingly, …

Expression Of Tumor-Associated Glycoprotein-72 (Tag-72) Antigen In Human Prostatic Adenocarcinomas., Dev Karan, Sonny L. Johansson, Ming-Fong Lin, Surinder K. Batra

Journal Articles: Biochemistry & Molecular Biology

Tumor-specific antigens are usually defined by monoclonal antibodies (MAbs) and can play critical roles in the diagnosis and therapy of carcinomas. Despite advances in the understanding of the molecular genetics of human prostate carcinomas, therapeutic approaches require that tumor-specific markers, preferably on the cell surface, should be defined. In this study, we examined the expression of an oncofetal antigen tumor-associated glycoprotein-72 (TAG-72) in prostatic adenocarcinomas with a Gleason grade of six or higher. Using a second generation MAb CC49 against TAG-72, immunoreactivity was detected in 88% (29/33) of the prostatic cancer tissues. Occasionally, the benign epithelium showed a very faint …

The Epidermal Growth Factor Receptor From Prostate Cells Is Dephosphorylated By A Prostate-Specific Phosphotyrosyl Phosphatase., Ming-Fong Lin, Gail M. Clinton

Journal Articles: Biochemistry & Molecular Biology

Human prostatic acid phosphatase (PAcP) has been found to have phosphotyrosyl-protein phosphatase activity (H. C. Li, J. Chernoff, L. B. Chen, and A. Kirschonbaun, Eur. J. Biochem. 138:45-51, 1984; M.-F. Lin and G. M. Clinton, Biochem. J. 235:351-357, 1986) and has been suggested to negatively regulate phosphotyrosine levels, at least in part, by inhibition of tyrosine protein kinase activity (M.-F. Lin and G. M. Clinton, Adv. Protein Phosphatases 4:199-228, 1987; M.-F. Lin, C. L. Lee, and G. M. Clinton, Mol. Cell. Biol. 6:4753-4757, 1986). We investigated the molecular interaction of PAcP with a specific tyrosine kinase, the epidermal growth factor …

Tyrosyl Kinase Activity Is Inversely Related To Prostatic Acid Phosphatase Activity In Two Human Prostate Carcinoma Cell Lines., Ming-Fong Lin, Ching-Li Lee, Gail M. Clinton

Journal Articles: Biochemistry & Molecular Biology

Alterations in prostatic acid phosphatase (PAcP), a phosphotyrosyl phosphatase, corresponded to changes in overall tyrosyl kinase activity. PAcP added to extracts of prostate carcinoma cells with a low endogenous level of PAcP activity and elevated tyrosyl kinase activity decreased the tyrosyl kinase activity. On the other hand, when PAcP activity was decreased by the addition of androgens to cells, there was a corresponding increase in tyrosyl kinase activity.