11 months ago

2006 Scientific Report

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Van Andel Research

Van Andel Research Institute | Scientific Report Role of integrins and RTKs in prostate epithelial cell survival Increased cell survival and resistance to cell death is a prerequisite for tumorigenesis. Several reports have suggested that the signaling pathways that regulate cell survival in normal prostate epithelial cells are different from the pathways regulating cell survival in prostate tumor cells. How integrin engagement of different ECMs regulates survival pathways in normal and tumor cells is not known. We have recently demonstrated that integrin-induced activation of EGFR in primary prostate epithelial cells is required for cell survival on laminin 5. The ability of EGFR to support integrin-mediated cell survival on laminin is mediated through α3β1 integrin and requires signaling downstream to Erk. Inhibition of integrin-induced EGFR/Erk signaling results in decreased expression of the anti-apoptosis regulator Bcl-XL and increased expression of the pro-apoptosis regulator BimEL (Fig. 1). Inhibition of Bcl-XL expression with siRNA induces cell death. However, despite this apparent regulation of the apoptotic machinery, cell death mediated by inhibition of EGFR or Bcl-XL occurs through a non–cytochrome C, non-caspase-dependent mechanism. Future studies in our lab are aimed at deciphering this cell death pathway. In contrast to the normal cells, EGFR and Erk are not activated by integrins in the metastatic PC3 prostate tumor cell line. Accordingly, integrin-mediated survival of PC3 cells does not depend on EGFR or Erk, but is instead dependent on PI-3K. Integrin-mediated survival through PI-3K in PC3 cells controls phosphorylation of the pro-apoptosis regulator Bad, which when blocked induces classic apoptotic cell death (Fig. 1). We will be testing additional prostate tumor cells lines to determine if this switch in matrix-mediated survival pathways is universally observed. 44 Figure 1. Laminin-mediated survival signaling pathways in normal and prostate tumor cells. Figure 1. Role of androgen receptor in integrin-mediated survival All primary and metastatic prostate cancers express the intracellular steroid receptor for androgen, AR. In the normal gland, the AR-expressing cells do not interact with the basement membrane; however, all AR-expressing tumor cells do adhere to the ECM in the basement membrane. In normal cells, AR expression suppresses growth and promotes differentiation, but in tumor cells AR expression promotes cell growth and is required for cell survival. The mechanisms that lead to the switch from growth inhibition and differentiation to growth promotion and survival are unknown. Our hypothesis is that adhesion to the ECM by the tumor cells is responsible for driving the switch in AR function. When prostate tumor cells are placed in culture, they lose expression of AR. The reason for this is not clear, but it may have something to do with loss of the appropriate ECM-containing basement membrane. When we introduce AR into prostate tumor cells, it actually suppresses their growth and induces cell death. However, if we place the AR-expressing tumor cells on laminin (the ECM found in tumors) and block the PI-3K signaling pathway, these cells do not die. The mechanisms responsible for this change in survival are unknown. Future work in the lab will be aimed at determining which cell survival pathways are activated by integrins.

VARI | 2006 Role of CD82 and integrin signaling in prostate cancer metastasis Prostate cancer death is due to the development of metastatic disease, which is difficult to control. The mechanisms involved in progression to metastatic disease are not understood. One approach that we are taking is to characterize genes that are specifically associated with metastatic prostate cancer. CD82/KAI1 is a metastasis suppressor gene whose expression is specifically lost in metastatic cancer, but not in primary tumors. Interestingly, CD82/KAI1 is known to associate with both integrins and RTKs. Our goal has been to determine how loss of CD82/KAI1 expression promotes metastasis by studying the role of CD82/KAI1 in integrin and RTK crosstalk. We have found that reexpression of CD82/KAI1 in metastatic tumor cells suppresses laminin-specific migration and invasion; integrininduced c-Met receptor and Src activation; and activation of the Src substrates Cas and FAK. Inhibition of either c-Met or Src also suppresses laminin-dependent invasion. In addition, CD82 suppresses HGF-induced activation of c-Met; however, if c-Met is overexpressed at sufficiently high levels, CD82 can no longer inhibit that activation. Thus, these data indicate that CD82/KAI1 normally acts to regulate both integrin- and HGF-mediated signaling to c-Met such that upon CD82 loss in tumor cells, signaling through c-Met is increased, leading to increased invasion. We are currently determining the mechanism by which CD82/KAI1 down-regulates c-Met signaling. So far our investigations indicate that c-Met and CD82 do not directly interact, and CD82 may act to suppress c-Met signaling by dispersing c-Met aggregates present on metastatic tumor cells into monomers and thus blocking signaling (Fig. 2). T = 0 T = 1h T = 4h Vector 45 Figure 2. CD82 reexpression in metastatic prostate cancer cells alters the distribution of c-Met (orange) on the surface of cells. CD82 Figure 2. We have also initiated several mouse studies to demonstrate the importance of CD82 in regulating metastasis in vivo. Using orthotopic injection of wild-type or CD82-expressing metastatic prostate tumor cells directly into the prostate, we found that CD82 also suppresses metastasis in vivo. We are continuing these studies using HGF-overexpressing mice and we hope to demonstrate that CD82 expression in metastatic prostate cells will suppress HGF-induced tumor formation in vivo. This will provide support for the role of CD82 in regulating c-Met function in vivo.

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