The influence of valproic acid and the role of cyclin D2 in prostate cancer

Prostate cancer (PCa) is the most frequently new diagnosed cancer and the third leading cause of cancer-related deaths in men in Germany. Studies on possible agents for PCa therapy are indispensable since treatment options for PCa are often associated with severe side effects. In the present study, the histone deacetylase inhibitor valproic acid (VPA) was investigated as a putative candidate for the treatment of prostate cancer. The expression of previously identified candidate genes, which are associated with PCa and exhibit a deregulated expression pattern in the murine PCa cells 2E upon VPA treatment, was analyzed by quantitative real-time PCR in prostate tumor tissue and prostate tissue of VPA-treated TRAMP mice. The molecular effects observed by in vitro VPA treatment of 2E cells could be partially confirmed in prostate tumor tissue and to a greater extend in prostate tissue of in vivo VPA-treated TRAMP mice. Furthermore, the influence of in vitro and in vivo VPA treatment on tumor- and tumor cell angiogenesis as well as on lymphangiogenesis was investigated by expression analyses of angiogenesis and lymphangiogenesis markers, including Vegfa, Vegfc, Ang1, Flt-1/Vegfr1, Kdr/Vegfr2, Tie-1, the soluble Vegfr2 receptor (sVegfr2) and Pecam-1. In murine PCa 2E cells, the expression Vegfa, Vegfc, Ang1, Tie-1 and Pecam-1 was decreased in a concentration- and partially time-dependent manner upon VPA treatment. In vivo VPA treatment also reduced the expression of several angiogenesis markers, as observed by statistically significant reduced Flt-1/Vegfr1 expression in prostate tissue and Ang1, Vegfa, Tie-1 and Kdr/Vegfr2 expression in prostate tumor tissue. These results indicate, that VPA treatment has a greater effect on the downregulation of angiogenesis markers in more advanced PCa than in very early tumor stages. The candidate gene cyclin D2 was studied in more detail since it was proposed to have a special function in PCa, possibly acting as a tumor suppressor. To elucidate the role of cyclin D2 in PCa cells, functional experiments of human PCa cells overexpressing cyclin D2 and of murine fibroblast cells with reduced cyclin D2 expression were conducted. Contrary to the expectation, functional analysis of human PCa cells with a transient, stable or inducible cyclin D2 overexpression revealed that these cells exhibited a similar or slightly increased proliferation rate as compared to controls. PC-3 and LNCaP cells with stable cyclin D2 overexpression had an increased migration rate as compared to controls. These data rather indicate that cyclin D2 is not a tumor suppressor in PCa. Functional studies of NIH/3T3 cells transfected with cyclin D2-specific siRNAs showed that these cells had an increased migration rate as compared to luciferase control transfected cells, indicating that cyclin D2 might have anti-tumorigenic capacities, nonetheless. Thus far, the hypothesis that cyclin D2 could act as a tumor suppressor in PCa remains unsolved