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- Title
- The Role of Type-I Interferon in Limiting Spread and Killing of an Oncolytic RNA Virus in Prostate Cells.
- Creator
-
Kedarinath, Kritika, Parks, Griffith, Chakrabarti, Ratna, Altomare, Deborah, University of Central Florida
- Abstract / Description
-
Prostate cancer is the second most prevalent cancer amongst men and there is an urgent need to address viable therapeutic options for its treatment. Development of viruses which target and kill cancer cells has gained momentum due to the first FDA approved oncolytic virus for treating human cancer patients. Our previous work with the RNA virus, Parainfluenza Virus 5 (PIV5), has led to the generation of mutants that are potential candidates for oncolytic viruses: 1) the hyperfusogenic (P/V/F)...
Show moreProstate cancer is the second most prevalent cancer amongst men and there is an urgent need to address viable therapeutic options for its treatment. Development of viruses which target and kill cancer cells has gained momentum due to the first FDA approved oncolytic virus for treating human cancer patients. Our previous work with the RNA virus, Parainfluenza Virus 5 (PIV5), has led to the generation of mutants that are potential candidates for oncolytic viruses: 1) the hyperfusogenic (P/V/F) mutant has a mutated P/V and fusion gene which activates anti-viral responses and causes massive cell-cell fusion respectively, and 2) the Leader mutant has a mutated viral genomic promoter which kills cells due to overactive viral gene expression. The P/V/F mutant has shown effectiveness in reducing prostate tumor burden in a mouse model system, however, the specificity of these viruses is unclear, i.e. targeting cancerous prostate cells while leaving uninvolved cells unaffected. In this study, we addressed how these PIV5 mutants replicate in and killed tumor versus benign human prostate cells. Flow cytometry demonstrated that the mutants are able to infect and replicate in prostate tumor cells (22Rv1), resulting in effective cell killing. However, these mutants showed highly restricted spread in benign prostatic hyperplasia cells (BPH-1). Upon further exploration, it was determined that the restriction observed in the BPH-1 cells is due to the induction and signaling of type-I Interferon (IFN). This was confirmed upon treatment with an IFN-? neutralizing antibody, which relieved restricted spread of mutants in benign cells. BPH-1 cells infected with the mutants also showed upregulation of key anti-viral, IFN-induced genes such as TLR3, IFIT1, and OAS2. Upon characterization of the mutant viruses in an additional metastatic prostate cancer cell line (C4-2B), a restriction in viral spread was observed. The restricted spread did not correlate with production of high levels of type-I IFN, suggesting that other cytokines or intracellular factors can limit replication in tumor cells. Therefore, these studies lay the groundwork for further improving the specificity of oncolytic PIV5 mutants by exploiting type-I IFN pathways as well as other anti-viral factors.
Show less - Date Issued
- 2016
- Identifier
- CFE0006468, ucf:51445
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0006468
- Title
- MECHANISM OF ACTION AND REGULATION OF MEMBRANE SERINE PROTEASE PROSTASIN IN THE PROSTATE AND PROSTATE CANCER.
- Creator
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Chen, Mengqian, Chai, Karl, University of Central Florida
- Abstract / Description
-
The glycosylphosphatidylinositol (GPI)-anchored serine protease prostasin (PRSS8) is expressed at the apical membrane surface of epithelial cells and acts as a suppressor of tumor invasion when re-expressed in highly invasive human prostate and breast cancer cell lines. To better understand the molecular mechanisms underlying the anti-invasion phenotype associated with prostasin re-expression in prostate cancer cells, we expressed wild-type human prostasin or a serine active-site mutant...
Show moreThe glycosylphosphatidylinositol (GPI)-anchored serine protease prostasin (PRSS8) is expressed at the apical membrane surface of epithelial cells and acts as a suppressor of tumor invasion when re-expressed in highly invasive human prostate and breast cancer cell lines. To better understand the molecular mechanisms underlying the anti-invasion phenotype associated with prostasin re-expression in prostate cancer cells, we expressed wild-type human prostasin or a serine active-site mutant prostasin in the PC-3 human prostate carcinoma cells. Molecular changes were measured at the mRNA and the protein levels. The expression of several invasion-promoting molecules is regulated by prostasin re-expression, mediated by a protein-level down-regulation of the epidermal growth factor receptor (EGFR). As a result, the cellular response to EGF was reduced as shown by the down-regulation of EGF-stimulated Erk1/2 phosphorylation. The expression of Slug, urokinase-type plasminogen activator (uPA), urokinase-type plasminogen activator receptor (uPAR), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and granulocyte-macrophage colony stimulating factor (GM-CSF) was also down-regulated by prostasin re-expression in the PC-3 cells. Co-expression of prostasin and its activating protease matriptase with EGFR in FT-293 cells induces an apparent proteolytic cleavage of the EGFR in the extracellular domain at two specific sites, generating two N-terminally truncated EGFR fragments, named EGFR135 and EGFR110. The EGFR110 is constitutively tyrosine-phosphorylated, and in its presence the phosphorylation of downstream signaling molecules including Erk1/2 and Akt is increased under serum-free conditions. Neither EGFR135 nor EGFR110 is responsive to EGF stimulation. Deletions of the EGFR extracellular domain (ECD) were generated to map the matriptase-prostasin cleavage sites. Two candidate sites were localized to regions AA1-273 and AA273-410. These data support a mechanism of action for the matriptase-prostasin epithelial extracellular serine protease activation cascade by proteolytically modulating the EGF-EGFR signaling. Prostasin gene expression is down-regulated in high-grade and hormone-refractory prostate cancers. We investigated the mechanisms by which androgens regulate prostasin expression in the prostate and prostate cancer. We treated the LNCaP human prostate cancer cells with dihydrotestosterone (DHT) and measured the mRNA expression of prostasin and potential transcription regulators of prostasin predicted by interrogation of the prostasin gene promoter sequence. Prostasin mRNA expression in the LNCaP cells was not responsive to DHT treatment. DHT marginally up-regulated mRNA expression of SREBP-1c, SREBP-2, and SNAIL, but not SREBP-1a, while dramatically increased SLUG mRNA expression, in a dose-dependent manner. Co-transfection of a prostasin promoter-reporter and SREBP cDNA in HEK-293 cells resulted in stimulation of the promoter activity at ~2 fold by SREBP-1c, and up to 6 fold by SREBP-2; while co-transfection with SNAIL or SLUG cDNA resulted in repression of the promoter activity to 43% or 59%, respectively. Co-transfection of the SLUG cDNA negated SREBP-2 s stimulation of the prostasin promoter in a dose-dependent manner. Transfection of an SREBP-2 cDNA in HEK-293 and DU-145 cells resulted in up-regulation of the endogenously expressed prostasin while transfection of a SLUG cDNA in the LNCaP cells repressed prostasin expression. Multiple SREBP-2 binding sites, known as sterol regulatory elements (SRE s), were identified at positions -897, -538, +8, +71, and +98 (named SRE-897, SRE-538, SRE+8, SRE+71, and SRE+98) in the human prostasin gene promoter. Mutagenesis of the five SRE s was carried out to evaluate their roles in SREBP-2 up-regulation of prostasin. SRE+98, a novel functional sterol regulatory element was found to be the major site for the stimulatory response of prostasin gene expression to SREBP-2. CONCLUSIONS: Prostasin regulates the expression of several invasion-promoting molecules in prostate cancer cells by down-modulating the EGF-EGFR signaling pathway. Active prostasin induces proteolytic cleavage in the EGFR ECD at two specific sites. One of the N-terminally truncated EGFR, the EGFR110 is auto-phosphorylated along with increased phosphorylation of downstream signaling molecules. The effect of the androgen DHT on prostasin expression in prostate cells is mediated via SREBP s, which stimulate the promoter, and Slug, which represses the promoter. Slug is up-regulated by DHT and EGF, providing a molecular mechanism by which epithelial cell-specific genes are silenced during prostate cancer development and progression.
Show less - Date Issued
- 2007
- Identifier
- CFE0001782, ucf:47257
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0001782
- Title
- IDENTIFICATION OF SMALL MOLECULES THAT INHIBIT PROSTATE CANCER CELL PROLIFERATION.
- Creator
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Zelaya, Rainel, Chakrabarti, Ratna, University of Central Florida
- Abstract / Description
-
Prostate cancer is the second most often diagnosed cancer and internationally the sixth foremost cause of cancer death in males, as of 2011. Within the United States it is the most common form of cancer in men with 186,000 new cases and with an overall 28,600 deaths in 2008, and it is the second leading kind of cancer-related death in men. The widespread threat that prostate cancer poses against men across the globe cannot be understated, and its initiation and progression must be understood...
Show moreProstate cancer is the second most often diagnosed cancer and internationally the sixth foremost cause of cancer death in males, as of 2011. Within the United States it is the most common form of cancer in men with 186,000 new cases and with an overall 28,600 deaths in 2008, and it is the second leading kind of cancer-related death in men. The widespread threat that prostate cancer poses against men across the globe cannot be understated, and its initiation and progression must be understood in order to truly comprehend its implicated risks and possible forms of treatment. As its name implies, prostate cancer is a form of cancer that develops in the prostate gland located in the male reproductive system. Its progress starts when standard semen-secreting prostate gland cells mutate into cancer cells. Although its developments may start at the prostate gland, cancer cells may metastasize to other parts of the body through circulation systems such as the lymph nodes. The main sites of metastasis for prostate cancer include the adrenal gland,the bones, the liver and the lungs. Although there are treatments available for prostate cancer, there is no definitive cure. The primary goal of this project was to find an alternative form of treatment, which is what will be necessary to combat this cancer.
Show less - Date Issued
- 2014
- Identifier
- CFH0004595, ucf:45228
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH0004595
- Title
- OUTCOMES ASSESSMENT WITHIN PROSTATE CANCER INTERVENTIONS FOR COUPLES: A NARRATIVE REVIEW.
- Creator
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Siguenza, Andrea M, Rovito, Michael J., University of Central Florida
- Abstract / Description
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Current cancer research is beginning to address the psychosocial implications of a prostate cancer (PCa) diagnosis from not just the perspective of quality of life of the patient, but of his partner as well. Such inquiries have created novel intervention programs aiming to alleviate the adverse side effects that a PCa diagnosis may inflict on the couple. Assessing efficacy of couple-based interventions, however, has been a difficult task due to the lack of homogeneity between studies...
Show moreCurrent cancer research is beginning to address the psychosocial implications of a prostate cancer (PCa) diagnosis from not just the perspective of quality of life of the patient, but of his partner as well. Such inquiries have created novel intervention programs aiming to alleviate the adverse side effects that a PCa diagnosis may inflict on the couple. Assessing efficacy of couple-based interventions, however, has been a difficult task due to the lack of homogeneity between studies regarding the operationalization process of primary outcome variables, as well as the instruments being used to measure them. This thesis, in response, aims to provide a detailed assessment of how previous interventions operationalized their targeted variables, the reported psychometric analysis of the instruments of measurement, and which instruments yielded statistically significant results. A narrative review was conducted using a database search strategy to collect articles regarding couple-based interventions that focused on outcomes related to PCa diagnosis, treatment, and survivorship. Out of the ten articles that passed the screening method, forty-two outcomes were identified, ranging from physical, social, and mental well-being of the couple, to the impact PCa had on their relationship quality. The outcomes were grouped into eight categories: quality of life, appraisal of PCa outcomes, sexual/physical well-being, relationship assessment, coping, mental health, knowledge, and distress. Various scales were used to measure similar outcomes with some articles failing to report on the psychometric properties of their chosen instruments. This assessment aims to provide future researchers with an indication as to what outcomes have been previously targeted and their corresponding methods of operationalization, categorization, and analysis. The multitude of assessed outcomes, the lack of uniformity on best practices in PCa couple intervention research, and the general failure to report on reliability and validity of measures may serve as significant barriers to producing high-quality evidence that can inform the development of future research and practice. This review provides the research community an aid in the development of behavioral interventions, and potentially, practice, via offering recommendations on certain outcomes that remain underreported within interventions. It is the ultimate aim of this project to assist in fostering a true public health for all.
Show less - Date Issued
- 2016
- Identifier
- CFH2000118, ucf:45977
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH2000118
- Title
- SCREENING FOR ANTICANCER AGENTS TO INHIBIT MITOTIC KINASES AND PROLIFERATION OF METASTATIC PROSTATE CANCER CELLS.
- Creator
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Nguyen, Khoa, Chakrabarti, Ratna, University of Central Florida
- Abstract / Description
-
Current treatments for prostate cancer (PCa) are marred with high relapse frequency and development of progressively aggressive cancers; developing new treatment options for PCa remains crucial. In this project, a series of synthetic compounds based on natural products will be screened to identify inhibitors for Aurora-A kinase (Aur-A). Aur-A facilitates centrosome separation and bipolar spindle formation during mitosis. Aur-A is overexpressed in metastatic PCa cells, and is a good candidate...
Show moreCurrent treatments for prostate cancer (PCa) are marred with high relapse frequency and development of progressively aggressive cancers; developing new treatment options for PCa remains crucial. In this project, a series of synthetic compounds based on natural products will be screened to identify inhibitors for Aurora-A kinase (Aur-A). Aur-A facilitates centrosome separation and bipolar spindle formation during mitosis. Aur-A is overexpressed in metastatic PCa cells, and is a good candidate for targeted therapies. Compound libraries are designed using natural compounds that contain simple structural elements as starting points for developing drug like libraries. High-throughput screening of these libraries will be used to identify potent antimitotic agents that selectively affect cancer cells but not normal cells. A combination of in vitro protein assays � quantifying protein activity � cell-based assays � measuring cell growth and proliferation � and cell-reporter assays � to determine which metabolic pathway the compound affects � were used to identify potential inhibitors. Through these methods, we have identified several compounds, with special consideration to thiazole piperazine compounds, to successfully inhibit proliferation of metastatic PCa cells.
Show less - Date Issued
- 2016
- Identifier
- CFH2000103, ucf:45549
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH2000103
- Title
- Variations in Health Services Utilization by Patients with Prostate Cancer.
- Creator
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McKee, Roberta, Wan, Thomas, Martin, Lawrence, Zhang, Ning, Sivo, Stephen, University of Central Florida
- Abstract / Description
-
Among men living in the United States, prostate cancer is the second leading cause of cancer death, and, excluding skin cancers, it is the cancer diagnosed most frequently. While incidence and mortality rates have been declining, the American Cancer Society estimated that there were 220,800 men diagnosed with prostate cancer and more than 27,500 prostate cancer deaths in 2015. Various patient-level and community-level factors have been shown to influence the differential patterns of diagnosis...
Show moreAmong men living in the United States, prostate cancer is the second leading cause of cancer death, and, excluding skin cancers, it is the cancer diagnosed most frequently. While incidence and mortality rates have been declining, the American Cancer Society estimated that there were 220,800 men diagnosed with prostate cancer and more than 27,500 prostate cancer deaths in 2015. Various patient-level and community-level factors have been shown to influence the differential patterns of diagnosis, care, and outcomes for men with prostate cancer. Detailed information regarding the utilization of health services by prostate cancer patients, particularly those with higher propensity for health services use, could be used to inform efforts intended to improve the coordination and delivery of care to work towards the elimination of disparities. The purpose of the study is to facilitate a better understanding of the determinants of health services utilization by older males with prostate cancer in the United States by examining the relative influence and interaction effects of factors characterizing individual patients and their county of residence. Andersen's behavioral model of health services utilization is used as a framework to guide this study. A cross-sectional design is used to analyze administrative claims data from the 2008 Medicare Provider Analysis Review (MEDPAR) file (n=5,754). County-level data from Area Health Resources File (ARHF) are merged to include the community and contextual characteristics. American Hospital Association (AHA) annual survey data are also used to examine the importance of hospital attributes in a subset analysis (n=555). A two-stage approach is used for analyzing the data. First, several social and demographic variables are included in automatic interaction detector (AID) analysis to identify relatively homogenous subgroups of patients with similar service utilization patterns for emergency room visits and hospital length of stay. Second, regression analysis is performed in the full dataset including all patients, and in each subgroup to determine the amount of variance explained by predictor variables categorized as predisposing, enabling, and need-for-care factors. Hierarchical logistic regression is performed to analyze the variability in emergency room use, and hierarchical multiple regression is performed to analyze the variability in hospital length of stay. The results show that the need-for-care factors are dominant predictors of service use. However, the relative importance of the predictor variables varies by subgroups of prostate cancer patients identified in the initial AID analysis. The findings lend some support of the use of an integrated approach to examine the personal and social determinants of health services utilization by prostate cancer patients enrolled in the U.S. Medicare program. The theoretical framework and analytic approach employed in this study make it possible to obtain an in-depth understanding of the influential factors associated with emergency room use and length of stay for all-cause hospitalizations, which can be used to inform future research and efforts aimed at developing targeted interventions to improve the coordinated care and to reduce health disparities among Medicare beneficiaries with prostate cancer.
Show less - Date Issued
- 2016
- Identifier
- CFE0006352, ucf:51523
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0006352
- Title
- The Influence of Stigma on Quality of Life and Relationship Satisfaction for Prostate Cancer Survivors and Their Partners.
- Creator
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Wood, Andrew, Barden, Sejal, Daire, Andrew, Lambie, Glenn, Munyon, Matthew, Conley, Abigail, University of Central Florida
- Abstract / Description
-
The purpose of this study was to examine the relationships between stigma, quality of life (QoL), and relationships satisfaction for prostate cancer (PCa) survivors and their intimate and/or romantic partners. The investigator tested a theoretical model that stigma (as measured by the Social Impact Scale [SIS; Fife (&) Wright, 2000]) influenced QoL (as measured by the Functional Assessment of Cancer Therapy (-) Prostate [FACT-P; Esper et al., 1997] and the Functional Assessment of Cancer...
Show moreThe purpose of this study was to examine the relationships between stigma, quality of life (QoL), and relationships satisfaction for prostate cancer (PCa) survivors and their intimate and/or romantic partners. The investigator tested a theoretical model that stigma (as measured by the Social Impact Scale [SIS; Fife (&) Wright, 2000]) influenced QoL (as measured by the Functional Assessment of Cancer Therapy (-) Prostate [FACT-P; Esper et al., 1997] and the Functional Assessment of Cancer Therapy (-) General Population [FACT-GP; Cella et al., 1993]) and relationship satisfaction (as measured by the Couples Satisfaction Index [CSI; Funk (&) Rogge, 2007]) for both PCa survivors and their partners (N = 72 couples). The investigator hypothesized that stigma would have a negative influence on both QoL and relationship satisfaction. Further, exploratory research questions pertained to the influence of race on stigma, QoL, and relationship satisfaction, as well as examining difference in experiences of stigma based on demographic variables (e.g., age and income).The results of the structural equation model analyses identified that stigma negatively influenced QoL (R2 = .84, p (<) .05) and relationship satisfaction (R2 = .19, p (<) .05) for both PCa survivors and their partners. Race did not have statistically significant (p (>) .05) relationships with stigma, QoL, or relationship satisfaction and stigma was not found to be statistically different (p (>) .05) based on demographic variables. Implications of the results of the study include (a) practical implications for PCa survivors and their partners; (b) strategies for effective individual, group, and couples-based counseling; (c) need for counselor educators to prepare counselors to work with medically ill populations and cancer survivors; (d) PCa stigma instrument development; and (e) the necessity to examine research with couples in a dyadic fashion.
Show less - Date Issued
- 2015
- Identifier
- CFE0005742, ucf:50112
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0005742
- Title
- MICRORNA REGULATION OF PROSTATE CANCER DESENSITIZATION TO ANDROGEN RECEPTOR ANTAGONIST DRUGS DURING ANDROGEN DEPRIVATION THERAPY.
- Creator
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Lorch, Robert, Chakrabarti, Ratna, University of Central Florida
- Abstract / Description
-
The current standard treatment of prostate cancer by androgen deprivation therapy involves using drugs such as bicalutamide (Casodex) to antagonistically block androgen receptors that are normally present within prostate cells. Usually, the therapy is successful in the short run at limiting the growth of prostate cancer. However, in virtually all cases tumors begin to grow aggressively again after several months of treatment and new therapies must be started. The mechanism by which these...
Show moreThe current standard treatment of prostate cancer by androgen deprivation therapy involves using drugs such as bicalutamide (Casodex) to antagonistically block androgen receptors that are normally present within prostate cells. Usually, the therapy is successful in the short run at limiting the growth of prostate cancer. However, in virtually all cases tumors begin to grow aggressively again after several months of treatment and new therapies must be started. The mechanism by which these prostate cells transform from androgen sensitive to androgen independent and anti-androgen resistant is unclear. In this study, we investigated the role of microRNAs, small 15 to 18 nucleotide regulatory RNAs, in regulating the desensitization of prostate cancer cells to the androgen receptor antagonist drug bicalutamide. In order to identify significant microRNAs, quantitative PCR was used to obtain genome-wide microRNA expression levels of 885 human microRNAs at different timepoints for androgen sensitive LNCaP cancer cells treated with bicalutamide and for untreated control cells in tissue culture. Analysis of microRNA expression by clustering analysis and by statistical comparisons of treatment groups resulted in identification of 28 microRNAs that have altered expression in the progression process. In silico target prediction analysis was performed with the microRNAs shown to have altered expression, and a group of genes predicted to be under microRNA regulatory control during cancer progression to resistance was identified. A microRNA expression profile can be useful in developing more effective prognostic and therapeutic tools for prostate cancer.
Show less - Date Issued
- 2011
- Identifier
- CFH0003826, ucf:44740
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFH0003826
- Title
- LIM KINASE 1 MODULATES EXPRESSION OF MATRIX METALLOPROTEINASES AND ASSOCIATES WITH GAMMA-TUBULIN: DUAL ROLE IN INVASION AND MITOTIC PROCESSES.
- Creator
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Tapia, Tenekua, Chakrabarti, Ratna, University of Central Florida
- Abstract / Description
-
LIM kinase 1 (LIMK1) is a unique dual specificity serine/threonine kinase containing two N-terminal LIM domains in tandem, a PDZ domain and a C-terminal catalytic domain. LIMK1 is involved in modulation of actin cytoskeleton through inactivating phosphorylation of the ADF (actin depolymerization factor) family protein cofilin. Recent studies have shown that LIMK1 is upregulated in breast and prostate cancer cells and tissues, promotes metastasis in animals and induces acquisition of an...
Show moreLIM kinase 1 (LIMK1) is a unique dual specificity serine/threonine kinase containing two N-terminal LIM domains in tandem, a PDZ domain and a C-terminal catalytic domain. LIMK1 is involved in modulation of actin cytoskeleton through inactivating phosphorylation of the ADF (actin depolymerization factor) family protein cofilin. Recent studies have shown that LIMK1 is upregulated in breast and prostate cancer cells and tissues, promotes metastasis in animals and induces acquisition of an invasive phenotype when ectopically expressed in benign prostate epithelial (BPH) cells. Furthermore, overexpression of LIMK1 was associated with altered sub cellular localization of the membrane type 1 matrix metalloprotease (MT1-MMP). Matrix metalloproteases (MMPs) are a family of zinc dependant proteolytic enzymes that hydrolyze extra cellular matrix and cell surface molecules. A number of MMPs including MMP-2, MMP-9 and their activator MT1-MMP are over expressed in a variety of cancers including prostate cancer. The abundant expression of these enzymes contributes to changes in the tumor microenvironment, which facilitate degradation of the surrounding collagen matrix and migration of cells through the matrix defects. In this study, we show that MMPs are involved in LIMK1 induced invasion of otherwise non-invasive BPH cells. We also show that (a) the kinase activity of LIMK is not essential for the invasive behavior of the cells and (b) the absence of LIM domains significantly retards cell invasion. We have established transfected sub lines of BPH cells stably expressing 1) constitutively active LIMK1 (BPHLCA), 2) kinase dead LIMK1 (BPHLKD) and 3) only the kinase domain of LIMK1 (BPHLK) for our study. In vitro invasion assays revealed that LIMK1 induced invasion was inhibited by the MMP specific inhibitor, GM6001, and that cells expressing kinase-dead LIMK1 were equally invasive. Furthermore, BPH cells expressing LIMK1 mutants expressed higher amounts of MMP-2 and MMP-9. Substrate zymography revealed increased concentration of secreted MMP-2 and MMP-9 in the media of BPHLCA and BPHLK cells respectively compared to BPHV (vector control) cells. Quantitative RT-PCR also showed a ~10 fold increase in the steady state concentration of MMP-2 in BPHLCA cells compared to the control BPHLV cells. Expression of active LIMK1 stimulated cell-surface expression of MT1-MMP in BPHLCA cells as determined by flow cytometry. A modest increase in expression of MT1-MMP was noted in BPHLKD cells compared to BPHLK and BPHV cells. Immunoflourescence analysis indicated differential localization of MT1-MMP and LIMK1 in BPH cells expressing different mutants of LIMK1. Co-localization of LIMK1 and MT1-MMP in the plasma membrane and in the perinuclear region was also evident in these cells. Furthermore, here we provide evidence that suggests a functional role for phosphorylated (activated) LIMK1/2 (p-LIMK1/2) during mitosis through its association with γ-tubulin. Immunoflourescence analysis showed distinct co-localization of γ -tubulin and p-LIMK1/2 in the centrosomes during mitosis from early prophase to the beginning of telophase. No association was seen in the interphase or in late telophase. Phospho-LIMK1/2 was co-precipitated in immunoprecipitates of γ -tubulin using an anti- γ -tubulin antibody suggesting a physical association between these proteins in a complex. This finding reveals a novel role of LIMK1 in the mitotic process. In summary, our data suggests that MMPs are involved in LIMK1 induced invasion of prostate epithelial cells, and that this effect is mediated through altered expression and activation of specific MMPs. Furthermore, LIMK1 induced invasion is dependant on the presence of LIM domains more than the kinase activity. Finally, we show that phosphorylated LIMK1 and LIMK2 are involved in the mitotic process in a stage specific manner through its association with the centrosomal protein γ -tubulin. Because LIMK1 promotes invasion in vitro, regulates expression of MMPs, and is involved in mitotic processes, it is an attractive drug target for prostate cancer therapy.
Show less - Date Issued
- 2007
- Identifier
- CFE0001812, ucf:47361
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0001812
- Title
- Identification and Functional Characterization of a Long Non-coding RNA associated with Prostate Cancer.
- Creator
-
Hasan, Md Faqrul, Chakrabarti, Ratna, Zhao, Jihe, Zhang, Shaojie, University of Central Florida
- Abstract / Description
-
Prostate cancer is the most common cancer in men in the western world. Although early stage prostate cancer is treatable late stage, more specifically, metastatic and drug resistant prostate cancers are mostly incurable. The failure of current treatments obligates the research community to explore novel areas in prostate cancer biology and find better therapeutic targets. Emerging evidences show that non-coding RNAs specifically long non-coding RNAs (lncRNAs) play regulatory roles in various...
Show moreProstate cancer is the most common cancer in men in the western world. Although early stage prostate cancer is treatable late stage, more specifically, metastatic and drug resistant prostate cancers are mostly incurable. The failure of current treatments obligates the research community to explore novel areas in prostate cancer biology and find better therapeutic targets. Emerging evidences show that non-coding RNAs specifically long non-coding RNAs (lncRNAs) play regulatory roles in various cellular processes and are frequently dysregulated in cancer including prostate cancer. These aberrantly expressed lncRNAs mostly with unexplored genetic information may drive cancer progression. Previous studies done in our laboratory showed a tumor suppressor role of a cluster of small non-coding RNAs or microRNA (miRNA) miR-17-92a in PC-3 prostate cancer cells. To learn the underlying mechanism, transcriptome analysis with or without expression of miR-17-92a was conducted in our laboratory. RNA-sequencing data analysis identified reduced expression of a set of lncRNAs and oncogenes, and up regulation of several tumor suppressor genes upon expression of miR-17-92a cluster miRNAs. One of the down regulated intergenic lncRNAs, PAINT (Prostate Cancer Associated Intergenic Non-coding Transcript) (LINC00888), was selected for determining its functional role in prostate cancer. TCGA and GEO profiles analyses revealed up regulation of PAINT in prostate tumors with higher Gleason Scores, in highly aggressive metastatic prostate cancer cell lines, and upon androgen deprivation therapy of prostate cancer cells. This observation was supported by our studies on expression analysis of PAINT in prostate tumor tissues using RNA in-situ hybridization in tissue microarrays (TMA) containing tissues from different stages of prostate cancer and normal prostate tissues, which showed higher expression of PAINT in prostate cancer tissues compared to normal tissues. Furthermore, late stage (stage III and stage IV) prostate tumors showed significant overexpression of PAINT compared to early stage (stage II) prostate cancer tissues. We examined the functional relevance of PAINT in promoting tumor progression next using different prostate cancer cell lines. Silencing of PAINT using siRNAs showed decreased cell proliferation, reduced S-phase progression and activation of pro-apoptotic proteins PARP and Caspase-3. Silencing of PAINT also showed decreased cell migration and increased expression of the epithelial marker, E-cadherin while reduced expression of mesenchymal markers Slug and Vimentin. Ectopic expression of PAINT reversed the effects observed upon silencing of PAINT. Increased cell proliferation, cell cycle progression and cell migration were noted in prostate cancer cells overexpressing PAINT. Additionally, cancer promoting phenotype such as larger colony formation and higher expression of mesenchymal marker Slug, was detected upon overexpression of PAINT. Our study also determined the therapeutic benefit of inhibition of expression showing an increased sensitivity of metastatic prostate cancer cells to the chemotherapeutic agent docetaxel (DTX) and selective Aurora kinase inhibitor VX-680. Taken together, our study establishes an oncogenic function of PAINT, its clinical relevance as a marker for advanced stage prostate cancer and its potential as a therapeutic target for metastatic prostate cancer.
Show less - Date Issued
- 2019
- Identifier
- CFE0007466, ucf:52681
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0007466
- Title
- Involvement of miRNAs in the Development of Androgen Independent Prostate Cancer.
- Creator
-
Ottman, Richard, Chakrabarti, Ratna, Cole, Alexander, Khaled, Annette, Zervos, Antonis, University of Central Florida
- Abstract / Description
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Development of resistance to androgen deprivation therapy (ADT) is a major obstacle for the management of advanced prostate cancer. Therapies with androgen receptor (AR) antagonists and androgen withdrawal initially result in tumor regression but development of compensatory mechanisms including AR bypass signaling leads to tumor re-growth, independent of circulating androgens. The result is the emergence of castration resistant prostate cancer (CRPC), a highly morbid disease exhibiting...
Show moreDevelopment of resistance to androgen deprivation therapy (ADT) is a major obstacle for the management of advanced prostate cancer. Therapies with androgen receptor (AR) antagonists and androgen withdrawal initially result in tumor regression but development of compensatory mechanisms including AR bypass signaling leads to tumor re-growth, independent of circulating androgens. The result is the emergence of castration resistant prostate cancer (CRPC), a highly morbid disease exhibiting aberrant expression of many protein-coding and non-coding genes. Under the umbrella of non-coding RNAs is a class of small regulatory RNAs referred to as microRNAs (miRNAs). MicroRNAs are believed to function in the maintenance of cell homeostasis but are often differentially expressed in many different types of cancer including CRPC.In this study, the association of genome wide miRNA expression (1113 unique miRNAs) with development of resistance to ADT was determined. Androgen sensitive prostate cancer cells that progressed to ADT and AR antagonist Casodex (CDX) resistance upon androgen withdrawal and treatment with CDX were used. Validation of expression of a subset of 100 miRNAs led to identification of 43 miRNAs that are significantly altered during progression of cells to treatment resistance. A correlation of altered expression of 10 proteins targeted by some of these miRNAs in these cells was shown.Additionally, profiles of miRNA expressions in cancerous prostate tissues were created and compared with profiles of paired adjacent uninvolved areas of prostate tissue. Among the miRNAs identified from these analyses, a cluster of miRNAs, miR-17-92a, that is under-expressed in prostate tumors and in androgen independent prostate cancer cells was highlighted. The miR-17-92a cluster miRNAs are transcribed from a polycistronic transcription unit C13orf25 that generates six mature miRNAs: miR-17, miR-18a, miR-19a, miR-19b, miR-20a and miR-92a, and is commonly de-regulated in many cancers. In this research, the expression of miR-17-92a miRNAs was found to be reduced in cancerous prostate tissues when compared to uninvolved areas and also in aggressive prostate cancer cells. Restoration of expression of all members of miR-17-92a cluster showed decreased expression of cell cycle regulatory proteins cyclin D1 and SSH1; as well as LIMK1 and FGD4 of the RhoGTPase signaling pathway. Expression of miR-17-92a miRNAs caused decreased cell proliferation, reduced activation of AKT and MAP kinases, delayed tumorigenicity and reduced tumor growth in animals. Additionally, miR-17-92a miRNA expression inhibited EMT via reduced cell migration and expression of mesenchymal markers while elevating expression and surface localization of the epithelial marker e-cadherin. Expression of miR-17-92a miRNAs improved sensitivity of androgen dependent LNCaP104-S prostate cancer cells to the Androgen Receptor antagonist bicalutamide (CDX), AKT inhibitor MK-2206 2HCl, and docetaxel. Androgen refractory PC-3 cells also showed increased sensitivity to docetaxel, MK-2206 2HCl, and Aurora kinase inhibitor VX680 upon ectopic expression of miR-17-92a cluster miRNAs. In conclusion, dynamic alterations in miRNA expression occur early on during androgen deprivation therapy and androgen receptor blockade. The cumulative effect of these altered miRNA expression profiles is the temporal modulation of multiple signaling pathways promoting survival and acquisition of resistance. These early events are driving the transition to castration resistance and cannot be studied in already developed CRPC cell lines or tissues. Notably, these data demonstrate a tumor suppressor effect of miR-17-92a cluster miRNAs in prostate cancer cells and restoration of expression of these miRNAs has a therapeutic benefit for both androgen-dependent and -independent prostate cancer cells. Furthermore, these results can be used as a prognostic marker of cancers with a potential to be resistant to ADT.
Show less - Date Issued
- 2016
- Identifier
- CFE0006697, ucf:52866
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0006697
- Title
- Light Scattering Property of Gold Nanoparticles with Applications to Biomolecule Detection and Analysis.
- Creator
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Zheng, Tianyu, Huo, Qun, Zou, Shengli, Gesquiere, Andre, Kang, Hyeran, Zhai, Lei, University of Central Florida
- Abstract / Description
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Gold nanoparticles (AuNPs) have unique optical and chemical properties. Dynamic light scattering (DLS) is an analytical tool used routinely for nanoparticle size measurement. The combined use of AuNPs and DLS has led to a novel analytical assay technology called D2Dx (from diameter to diagnostics). Herein, my dissertation highlights the extended use of D2Dx for biomolecule detection and analysis. Under this general theme, Chapter 1 provides some background information of AuNPs, DLS, the...
Show moreGold nanoparticles (AuNPs) have unique optical and chemical properties. Dynamic light scattering (DLS) is an analytical tool used routinely for nanoparticle size measurement. The combined use of AuNPs and DLS has led to a novel analytical assay technology called D2Dx (from diameter to diagnostics). Herein, my dissertation highlights the extended use of D2Dx for biomolecule detection and analysis. Under this general theme, Chapter 1 provides some background information of AuNPs, DLS, the principle of D2Dx technique and its potential applications. Chapter 2 summarizes a study on the effect of AuNP concentrations and laser power on the hydrodynamic size measurement of AuNPs by DLS. This study demonstrated the multiple scattering effect on DLS analysis, and how to use the exceptionally high sensitivity of DLS in AuNP aggregate detection for bioassay design and development. Chapter 3 explores a cooperative interaction between AuNP and certain proteins in blood serum that are key to the immune system, leading to a novel diagnostic tool that can conveniently monitor the humoral immunity development from neonates to adults and detect active infections in animals. Chapter 4 reports an application of D2Dx technique for acute viral infection detection based on the active immune responses elicited from mouse models infected with influenza virus. Chapter 5 describes another application of D2Dx for prostate cancer detection. The D2Dx assay identifies prostate cancer patients from non-cancer controls with improved specificity and sensitivity than PSA test. Chapter 6 demonstrates the use of AuNPs and DLS for hydrodynamic size measurement of protein disulfide isomerase with two different conformations. Chapter 7 investigates the concentration-dependent self-assembling behavior of ribostamycin through its interaction with AuNPs in aqueous solution. Overall, this dissertation established several lines of applications of using AuNPs and DLS for biomolecular research and in vitro diagnostics.
Show less - Date Issued
- 2018
- Identifier
- CFE0007385, ucf:52056
- Format
- Document (PDF)
- PURL
- http://purl.flvc.org/ucf/fd/CFE0007385