[Show abstract][Hide abstract] ABSTRACT: Invasion and subsequent metastasis is the major cause of death from most cancers including prostate cancer. Herein we report on the potential tumor suppressive properties of Rab7, a GTPase that regulates trafficking of lysosomes. The movement of lysosomes to the cell surface in response to environmental cues increases the secretion of proteinases and cell invasion. We determined that Troglitazone and other members of the Thiazolidinedione family inhibit cell-surface directed lysosome trafficking and cathepsin B secretion through a Rab7-dependent mechanism. Moreover, Rab7 shRNA expressing cells were found to be more invasive in vitro and in vivo. Increased invasiveness was accompanied by elevated expression of the c-Met receptor and prolonged downstream signaling, thereby supporting a role for Rab7 as a mediator of signaling down-regulation. Taken together, these results suggested that Rab7 acts as a negative regulator of prostate tumor growth and invasion, providing further evidence for its potential as a tumor suppressor.
PLoS ONE 01/2014; 9(2):e87882. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Human prostate tumor vaccine and gene therapy trials using ex vivo methods to prime dendritic cells (DCs) with prostate specific membrane antigen (PSMA) have been somewhat successful, but to date the lengthy ex vivo manipulation of DCs has limited the widespread clinical utility of this approach. Our goal was to improve upon cancer vaccination with tumor antigens by delivering PSMA via a CD40-targeted adenovirus vector directly to DCs as an efficient means for activation and antigen presentation to T-cells. To test this approach, we developed a mouse model of prostate cancer by generating clonal derivatives of the mouse RM-1 prostate cancer cell line expressing human PSMA (RM-1-PSMA cells). To maximize antigen presentation in target cells, both MHC class I and TAP protein expression was induced in RM-1 cells by transduction with an Ad vector expressing interferon-gamma (Ad5-IFNγ). Administering DCs infected ex vivo with CD40-targeted Ad5-huPSMA, as well as direct intraperitoneal injection of the vector, resulted in high levels of tumor-specific CTL responses against RM-1-PSMA cells pretreated with Ad5-IFNγ as target cells. CD40 targeting significantly improved the therapeutic antitumor efficacy of Ad5-huPSMA encoding PSMA when combined with Ad5-IFNγ in the RM-1-PSMA model. These results suggest that a CD-targeted adenovirus delivering PSMA may be effective clinically for prostate cancer immunotherapy.
PLoS ONE 01/2012; 7(10):e46981. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The Tousled-like kinases (TLKs) function in processes of chromatin assembly, including replication, transcription, repair, and chromosome segregation. TLK1/1B interacts specifically with the chromatin assembly factor Asf1, a histone H3-H4 chaperone, and with Rad9, a protein involved in DNA repair, and these interactions are believed to be responsible for the action of TLKs in double-strand break repair and radioprotection.
Western blotting and RT-PCR were used to analyze the expression of TLK1, TLK1B, and TLK2 in a panel of prostate cancer (CaP) cell lines. The pattern of radiotolerance in the cell lines was analyzed in parallel. DU145 and PC-3 cells were also probed with assays utilizing transfected plasmids that could be cleaved in vivo with adeno-expressed HO nuclease to assess the potential contribution of TLK1/1B in DSB repair.
This is the first report of TLKs' expression in a panel of CaP cell lines and their relationship to radioresistance. Furthermore, expression of TLK1B in non-expressing PC-3 cells rendered them highly resistant to radiation, and conversely, knockdown to TLK1/1B in expressing DU145 reduced their radiotolerance.
TLKs appear to be intimately linked to the pattern of resistance to DNA damage, and specifically DSBs, a finding that was not reported before for any cell lines, and certainly not systematically for human prostate cell lines.
The Prostate 02/2011; 71(13):1367-73. · 3.84 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Core binding factor (CBF) is a transcription factor complex that plays roles in development, stem-cell homeostasis, and human disease. CBF is a heterodimer composed of one of three DNA-binding RUNX proteins plus the non-DNA-binding protein, CBFβ. Recent studies have showed that the RUNX factors exhibit complex expression patterns in prostate, breast, and ovarian cancers, and CBF has been implicated in the control of cancer-related genes. However, the biologic roles of CBF in solid tumors have not been fully elucidated. To test whether CBF is required for the malignant phenotype of various epithelial cancers, we used lentiviral delivery of CBFβ-specific shRNA to significantly decrease CBFβ expression in two prostate cancer cell lines (PPC1 and PC-3) and the SKOV-3 ovarian cancer cell line. We found that knockdown of CBFβ significantly inhibited anchorage independent growth of each cell line. Further, CBFβ knockdown in PPC1 cells suppressed xenograft tumor growth compared to controls. Mice injected with SKOV-3 ovarian cancer cells knocked-down for CBFβ exhibited a survival time similar to control mice. However, human cells recovered from the ascites fluid of these mice showed CBFβ expression levels similar to those from mice injected with control SKOV-3 cells, suggesting that CBFβ knockdown is incompatible with tumor cell growth. Gene expression profiling of CBFβ knockdown cells revealed significant changes in expression in genes involved in various developmental and cell signaling pathways. These data collectively suggest that CBFβ is required for malignancy in some human cancers.
Journal of Cellular Physiology 11/2010; 225(3):875-87. · 4.22 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The insulin-like growth factor (IGF) axis is associated intimately with prostate cancer (PCa) development, growth, survival and metastasis. In particular, increased levels of IGFBP-2 expression are associated with advanced PCa, bone metastasis, and the development of castrate resistant PCa. Previously, we reported that androgen treatment decreased intracellular and extracellular IGFBP-2 in the androgen sensitive (AS) PCa cell line, LNCaP. Nonetheless, the mechanism by which androgen treatment decreases expression of IGFBP-2 is not clear. Since elevated IGFBP-2 is associated with a variety of advanced cancers, including PCa, coupled with the fact that hormone ablation is the customary treatment modality for advanced PCa, a complete understanding of the influence of androgens on IGFBP-2 expression is essential. Androgen treatment initially increased steady state IGFBP-2 mRNA levels in LNCaP cells. Extended androgen treatment on LNCaP resulted in a time-dependent decrease in both steady state IGFBP-2 mRNA and protein. Polysomal mRNA analysis showed no difference in IGFBP-2 association with a given fraction; however, Q-PCR revealed less IGFBP-2 mRNA in each androgen-treated fraction. In addition, there was an overall decrease in polysome mRNA after androgen treatment. Extracellular proteolysis of IGFBP-2 was prevented in the presence of serine protease inhibitors. These data indicate that androgen acts via multiple levels to down-regulate IGFBP-2 in LNCaP PCa cells.
American Journal of Translational Research 01/2010; 2(2):200-8.