טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentInbar Michal
SubjectRegulation of Growth Hormone Binding Protein Generation
and Leptin Signaling in Human Prostate Cancer Cell
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DepartmentDepartment of Medicine
Supervisor Assistant Professor Ronnie Barkey
Full Thesis textFull thesis text - English Version


Abstract

Growth hormone binding protein (GHBP) is a major regulator of GH action. Therefore, it is reasonable to assume that GHBP plays a regulatory role in the cancer promoting effects of GH. This research studied the involvement of GHBP in PCa, by examining the involvement of several signaling pathways in GHBP shedding.

Treatment of CHO/hGHR control cells with phorbol ester (PMA) resulted in a 50% enhancement in GHBP generation, versus ≤3-fold by androgen-dependent LNCaP PCa cells. These results indicate involvement of protein kinase-C (PKC) in GHBP shedding. Indeed, a PKC inhibitor completely inhibited PMA-induced GHBP shedding from CHO/hGHR cells and caused partial inhibition in LNCaP cells.

Inhibitors of extracellular signal regulated kinase (ERK)1/2-MAPK (mitogen activated protein kinase), p38-MAPK and phosphatidyl-inositol-3-kinase caused a significant decrease in PMA-induced GHBP shedding in CHO/hGHR and LNCaP cells, indicating the involvement of these pathways in GHBP generation.

Serum induced a 7-fold increase in GHBP shedding from CHO/hGHR cells, which was significantly inhibited by pre-treatment with kinase inhibitors. These results concur with our findings that serum induced PMA shedding to detectable levels in LNCaP cells. These differential effects of serum provide a convincing basis for our research hypothesis, that cancer-induced local differences in serum composition may be responsible for differential regulation of GHBP shedding and reciprocally, that GHBP may play a regulatory role in PCa.

The cytokine hormone leptin is formed in and secreted by adipocytes and its levels are elevated in obesity. Obesity is a risk factor for many diseases including cancer. Leptin is a mitogen in many cell types and was shown to activate the Janus Kinase 2 (JAK2) and MAPK-ERK1/2 signaling pathways.

The second part of this study investigated the molecular effects of leptin in human PCa.

Following overexpression of leptin receptor (LR) and JAK2, leptin induced activation of JAK2, STAT3 and ERK1/2 in DU145, PC-3, PC-3/AR and LNCaP human PCa cells. JAK2 activation was more evident in AR-positive than AR-negative cells.

Co-transfection with kinase-negative (KN)-human epidermal growth factor receptor (HER)2-YFP cDNA, in addition to LR and JAK2, caused leptin-induced phospho-JAK2 to transactivate KN-HER2. This effect was more evident in AR-positive than AR-negative cells, suggesting that androgen-sensitive PCa, which overexpresses HER2, may become more reactive to cytokine action. Leptin also exerted long-term effects on KN-HER2 and MAPK phosphorylation and increased AR protein level in LNCaP cells.

Together, these results suggest that leptin may play an important role in cellular events leading to PCa progression.