Abstract The immune modulatory drug lenalidomide has shown promising anti-tumor activity in a clinical trial of patients with advanced hepatocellular carcinoma (HCC). The present study explored whether lenalidomide can enhance the anti-tumor activity of sorafenib, the standard molecular targeted therapy for HCC. An orthotopic liver cancer model was developed by injecting the BNL liver cancer cells (from ATCC) into the liver capsule of BALB/c mice. Mice were treated by sorafenib by daily gavage or lenalidomide by daily intra-peritoneal injection, and the anti-tumor efficacy of single-agent or combination treatment was measured by change in tumor volume and animal survival. The number of IFN-γ expressing T-cell sub-populations in tumor-infiltrating lymphocytes (TILs) and the number of IFN-γ expressing tumors-specific CD8+ T cells in splenocytes derived from tumor-implanted mice were measured by flow cytometry. Depletion of relevant T cell sub-populations or cytokines was done by co-administration of relevant antibodies with study drug treatment. TUNEL assay and immunohistochemical study on tumor tissues were done to measure apoptosis and to explore potential biomarkers (Ki67, CD31). Combination of sorafenib (5 mg/kg/day) and lenalidomide (50 mg/kg/day) produced significant synergistic anti-tumor efficacy in terms of tumor growth delay and animal survival. This synergistic effect was associated with a significant increase in IFN-γ expressing CD8+ lymphocytes in TILs and splenocytes. In addition, TILs derived from mice in the combination treatment group had higher number of granzyme- or perforin- expressing CD8+ T cells, compared with vehicle- or single-agent-treatment groups. Combination treatment was associated with a significant increase in apoptotic tumor cells and decrease in microvessel density in tumor tissue. Finally, the synergistic anti-tumor effect was abolished after CD8 depletion. The combination treatment was well tolerated except that leucopenia was common. Our data indicate that lenalidomide can enhance the anti-tumor effects of sorafenib in HCC through its immune modulatory effects and CD8+ T cells play an important role in the anti-tumor synergism. (supported by grants NSC 101-2321-B-002 -014 and NSC 101-2314-B-002-162) Citation Format: Chun-Jung Chang, Da-Liang Ou, Yi-Jang Lin, Ann-Lii Cheng, Chiun Hsu. Potential synergistic antitumor activity between sorafenib and lenalidomide in an orthotopic murine liver cancer model. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr B10.