Date of Award

6-1-2012

Degree Type

Dissertation

University or Center

Clark Atlanta University(CAU)

School

School of Arts and Sciences

Degree Name

Ph.D.

Department

Biology

First Advisor

Dr. Cimona V. Hinton

Second Advisor

Dr. Valerie Odero-Marah

Third Advisor

Dr. Ritu Aneja

Abstract

Inactivation of the tumor suppressor phosphatase and tensin liomolog deleted on chromosome 10 (PTEN) is heavily implicated in the tumorigenesis of prostate cancer. Conversely, the upregulation of the chemokine (CXC) receptor 4 (CXCR4) is associated with prostate cancer progression and metastasis. The inactivation of functional PTEN is typically due to genetic and epigenetic modulations, as well as active site oxidation by reactive oxygen species (ROS); likewise ROS upregulates CXCR4 expression. Furthermore, the pathways of CXCR4 and PTEN converge, leading to the promotion and regulation of turnorigenesis, respectively. Based on these observation we hypothesized that, PTEN inhibits CXCR4-mediated functions in prostate cancer cells. This hypothesis was investigated through the following aims: (1) To determine whether PTEN inhibits CXCR4-mediated functions in prostate cancer cells and, (2) To investigate a mechanism by which PTEN function is lost, and CXCR4 function is gained in prostate cancer cells. Upon screening advanced metastatic prostate cancer cell lines for PTEN, identified a loss of expression in PC3 and LNCaP cells, whereas DU145 expressed wildtype PTEN. All three cell lines were positive for surface expression of CXCR4. Reconstitution of PTEN induced a mesenchymal to epithelial-like morphological change and inhibited CXCR4-mediated migration and proliferation in PC3 cells. Downregulation of PTEN by siRNA enhanced the CXCR4-mediated migratory behavior of DU145 cells. By western blot analysis, we observed that PTEN inhibited basal Protein Kinase B (AKT) phosphorylation, but not extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation in PTEN expressing cells. Upon CXCR4 stimulation, PTEN inhibited ERK1/2 phosphorylation, but not phosphorylation of AKT. The CXCR4- mediated migration of PC3 cells was through the ERK1/2 pathway, as confirmed by chemical inhibitors. Based on these studies, we suggest that loss of PTEN permits CXCR4-mediated functions in prostate cancer cells, through the ERK1/2 pathway. We also observed that ROS increased expression of phosphorylated AKT (p -AKT) and phosphorylated ERK1/2 (p-ERK1/2) in DU145 and 22Rv1 prostate cancer cell lines, which was abrogated by the ROS scavenger N-acetyl cysteine (NAC). This study showed that PTEN, was oxidized by ROS accumulation. The oxidized form of PTEN became catalytically inactive, although the expression levels of PTEN remained consistent. Additionally, the ROS-mediated inactivation of PTEN enhanced hypoxia inducible factor lcL (HIF1CL) and CXCR4 expression, and invasion and migration of DU145 cells, in a CXCR4-dependent manner. Interestingly, at the same ROS concentration, 22Rv1 cells did not display mitogenic effects. HIFlo expression decreased in 22Rv1 cells and the cells became apoptotic, despite the increases in AKT and ERK1/2. Taken together, antagonizing CXCR4 and downstream signaling cascades may provide an efficient approach for treating patients with advanced prostate cancer.

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