which were more than two logs fold efficacious in infected mice compared with P

Both S1PR2 shRNA sequences greatly reduced Ad AC induced Akt activation, confirming a prominent role for S1PR2 signaling in the activation of Akt downstream of AC. As the observation that S1PR2 activates an oncogenic signaling pathway challenges the dogma on the role of S1PR2 in cancer Aurora Kinase Inhibitor cell signaling, we performed a proliferation experiment and found that the proliferation advantage of AC overexpressing prostate cancer cells is diminished by treatment with JTE013. Basal S1PR1?3 expression was evaluated in PPC1 and DU145, both of which had predominate S1PR2 mRNA with markedly less S1PR1 and 3. Further analysis revealed that S1PR2 mRNA is induced slightly, but significantly, upon AC expression, whereas the other ceramidases are not affected by AC expression, except for a reduction in ACER1 mRNA in PPC1. S1PRs are GPCRs known to stimulate Akt activation by activating Gi mediated stimulation of PI3K. Pertussis Skin infection toxin, which inactivates Gi, G0 and Gt, prevented AC induced Akt activation, and the Gi inhibitor NF023 abrogated AC induced Akt activation, suggesting a role for G proteins, specifically Gi, in AC induced Akt activation. Expressing PTEN in PPC1 cells antagonized AC induced Akt activation, and the PI3K inhibitor LY294002 effected dose dependent abrogation of pAkt, supporting an S1PR2, PI3K dependent mechanism. To test whether exogenous S1P works in the same way on these cell lines, we treated PPC1 and DU145 with 500 nM S1P for 2 h in the presence or absence of JTE013. JTE013 blocked S1P induced Akt activation in both cell lines, supporting the findings using AC expression to drive increased S1P signaling. BIX01294 AC promotes chemotherapy resistance, but confers sensitivity to Akt inhibition Cytotoxic chemotherapy depends, in part, on ceramide accumulation to cause cell death. PPC1 cells were subjected to a wide dose range of the cytotoxic chemotherapeutic agents Docetaxel, Gemcitabine and 50 Fluorouracil. PPC1 cells infected with Ad AC were found to be less sensitive to all the three compounds, reflected by an increased EC50. Conversely, AC overexpressing cells were more sensitive to inhibition of Akt with Akt inhibitor X, Perifosine or MK2206, with AC expressing cells being B30?40% more sensitive than Ad GFP infected cells. Proliferation in AC overexpressing cells is profoundly sensitive to Akt inhibition Akt signaling promotes cancer in numerous ways, including increased cell proliferation. To determine whether AC induced proliferation is Akt dependent, we evaluated prostate cancer cell proliferation in the presence of AktX and Perifosine. In DU145 ACEGFP cells stably expressing AC, we noted significantly more rapid cell proliferation compared with the vector control. Treatment with AktX and Perifosine both reduced proliferation in AC EGFP and EGFP cell lines.