Ansfection of the C/EBPa siRNAs but not the controlor C/EBPb siRNAs abrogated L803 mts induced E2F1 suppression. These data Rocuronium Zemuron indicate that C/EBPa protein is involved in GSK 3 inhibition induced E2F1 suppression in prostate cancer cells. DISCUSSION The goal of this study was to establish a proof of concept that suppression of GSK 3 activity leads to reduced tumor development and growth of prostate cancer in vivo. We used two animal models, subcutaneous xenograft and spontaneous TRAMP tumor. In mouse xenograft models, GSK 3 inhibition suppressed xenograft tumor development and growth. Again, with the TRAMP model, GSK 3 inhibitors reduced the tumor burden of cancerous prostate tissues in mice. These anti tumor effects were associated with impaired DNA synthesis and reduced cell proliferation as assessed by BrdU labeling and Ki 67 immunostaining. These data are in agreement with our previous findings that GSK 3 inhibitors reduced prostate cancer cell proliferation in vitro by blocking S phase gene expression, as well as other reports from different groups. In conjunction with other reports that GSK 3 inhibitors have therapeutic potential for human cancers through various mechanisms, our results provided additional support to the notion that GSK 3 inhibitors might be developed as new agents for prostate cancer intervention, particularly for late stage advanced diseases. Most importantly, in this study,wedocumented that GSK 3 inhibition resulted in accumulation of C/EBPa protein. Sequential analysis revealed that C/EBPa accumulation is responsible for GSK 3 inhibitorinduced suppression of E2F1 transactivation. Although most of previous studies regarding C/EBPa function are related to adipocyte differentiation, emerging data suggest that it might also act as a tumor suppressor. In prostate cancer cells, C/EBPa has been found to negatively regulate AR mediated gene expression and cell proliferation in prostate cancer cells.
In this study, we found that C/EBPa protein levels increased rapidly but the mRNA levels remained unchanged following GSK 3 inhibition, suggesting that GSK 3 inhibition reduced C/EBPa protein degradation. Since GSK 3 has been shown to phosphorylate C/EBPa and GSK 3 mediated phosphorylation usually Honokiol results in its substrate degradation via ubiquitin proteasome pathway, it is plausible that GSK 3 inhibition resulted in reduced C/EBPa phosphorylation and subsequently proteasome mediated degradation. On the other hand, a previous report showed that lithiuminduced C/EBPa accumulation in a mouse keratinocyte model was due to reduced proteasome dependent proteolytic degradation, but GSK 3’s role in C/EBPa stability was not clearly defined in their study. This inconsistency suggests a cell specific difference in C/EBPa regulation. Nonetheless, our data indicated C/EBPa accumulation is involved in GSK 3 inhibitioninduced cytostatic effect in prostate cancer cells, although the detailed mechanism of GSK 3 dependent regulation of C/EBPa protein stability requires further investigation. Currently, advanced prostate cancers after escaping androgen ablation therapy are virtually no means to cure. Therefore, agents that target unique cellular signal pathways such as GSK 3 might represent novel therapeutic targets for these patients.