We observed that 25% of the male ATX/IRS-1 mice developed HCC with a long latency of 15 months; single ATX and IRS-1 transgenic mice were tumor-free. The liver was examined for histologic changes and liver tissue and tumor lysates

were prepared for mRNA and protein expression by qRT-PCR and multiplex ELISAs respectively. Expression and phosphorylation of multiple signaling molecules were analyzed in the insulin/IGF-1/IRS-1/Ras/Raf/MAPK/Erk, and PI3K/Akt/GSK3, Wnt/β-catenin, and Notch signaling cascades. These are highly conserved signaling pathways that interact and crosstalk with each other, and found active in the majority of human HCC tumors. Results: In the ATX/IRS-1 liver, early activation of the insulin/IGF pathway as evidenced by increased phosphorylation of IgF-1 R and Erk was followed by activation of the Wnt/β-catenin and Notch Palbociclib cell line pathways in the form

of Wnt-3, FZD-7, CCND-1, TBX-3, Notch-1, JAG-1, and Hes-1 overexpression. In the ATX/IRS-1 -derived HCC tumors, upregulation of these pathways was further accompanied by highly expressed levels of IGF-2, Wnt-3, FZD-7, cyclin D-1, and Hes-1. Conclusions: These studies demonstrate that the insulin/IGF-1, Wnt/β-catenin, and Notch signaling cascades are activated in ATX/IRS double-transgenic murine model. This was the result of the synergistic action between the HBx and IRS-1 overexpression which linked these interacting signaling pathways. The timing of pathway activation to the relationship of histopathologic changes

in the liver from normal to dysplas-tic AZD9291 purchase hepatocytes to HCC Cetuximab ic50 strongly supports the hypothesis that upregulation of the insulin/IGF-1 pathway alone results primarily in accelerated hepatocyte proliferation, but the synergistic upregulation of all three signaling pathways is necessary and sufficient to initiate HCC tumor formation. Disclosures: The following people have nothing to disclose: Waihong Chung, Suzanne M. de la Monte, Miran Kim, Jack R. Wands Background: Interleukin 12 (IL-12) as one of the most potent Th1-cytokines has been used to improve dendritic cells (DC)-based immunotherapy of cancer. However, this approach failed to achieve clinical response in patients with hepatocellular carcinoma (HCC). In this study, an improved immunotherapy with DC engineered to express IL-12 was studied in murine subcutaneous HCC. Methods: Tumor-lysate pulsed DC were transduced with IL-1 2-encoding adenoviruses or cultivated with recombinant (r)IL-12. DC were injected intratumorally (i.t.), sub-cutaneously or intravenously at different stages of tumor-development. In a further step, immunotherapy with DC was combined with a daily oral treatment of sorafenib (30mg/kg body weight). The tumor environment was characterized using flow cytometry, Elisa and immunohistochemistry regarding t-cell recruitment and cytokine expression.