Abstract A010: Interrogation of checkpoint inhibitors in bioluminescent orthotopic syngeneic models of hepatocellular carcinoma
Authors: Papadopoulou Nektaria , Wainwright Louise , Lacey Vicky , Wrigley Jane , Wood Jamie , Malhi Sumanjeet , Woolley Louise , King Jason , Jiang Simon , Kumari Rajendra 
Published: August 2018

Abstract
Background: Hepatocellular carcinoma (HCC) is an aggressive disease with limited therapeutic treatment options. Checkpoint inhibitors are widely being evaluated in numerous cancers including HCC, but suitable preclinical models are required. Subcutaneous syngeneic models are widely used to evaluate the impact of immunotherapy on tumor growth and tumor-invading leucocytes (TILs). However, the tumor microenvironment of orthotopic models is more comparable to the patient as they form an organ-specific site, facilitate metastatic spread, and, in the case of syngeneic models, support immune and stromal component interactions. Bioluminescent imaging (BLI) enables noninvasive longitudinal monitoring of orthotopic tumor burden, allowing for optimal randomization, reduction of false positives, and continuous feedback supporting optimal treatment regimen mid-study. Here we describe the generation of bioluminescent variants of syngeneic hepatocellular carcinoma (HCC) cell lines and assess the impact of immune checkpoint therapy on orthotopic growth in the liver. Methods: Bioluminescent variants of syngeneic cell lines H22 and Hepa 1-6 were established by lentiviral transduction. Orthotopic models were established via direct implantation into the liver. Under anesthesia a small transverse incision was made below the xiphoid to expose the liver, and cells were injected in the upper left lobe of the liver. The liver was then carefully placed back into the abdominal cavity and sutured. Tumor growth was assessed by BLI twice weekly and at end stage (Spectrum CT; PerkinElmer). Response to sorafenib and immune checkpoint therapy was also evaluated and TIL infiltration was assessed by FACs analysis and IHC. Results: The success rate of tumor transplantation into the liver was 100% as confirmed by both in-life imaging and ex vivo imaging at termination. Hepa 1-6 appeared to grow within the parenchyma of the liver whereas H22 grew within the liver as well as ascites. Real-time quantification of tumor size in response to treatment with sorafenib and check point inhibitors was correlated with alpha-fetoprotein (AFP) and end-stage tumor burden. Differences in TIL infiltration as detected by FACs and IHC were observed as well as stromal and blood capillary infiltration. Conclusions: The growth of bioluminescent HCC syngeneic models Hepa 1-6 and H22 was established to enable the evaluation of standard-of-care agents such sorafenib as well as immunotherapeutic agents. The liver microenvironment influences the orthotopic tumor growth and TIL infiltration, which requires characterization by both FACS and IHC in order to understand the relevance and utilization of such models.
Citation Format: Nektaria Papadopoulou, Louise Wainwright, Vicky Lacey, Jane Wrigley, Jamie Wood, Sumanjeet Malhi, Louise Woolley, Jason King, Simon Jiang, Rajendra Kumari. Interrogation of checkpoint inhibitors in bioluminescent orthotopic syngeneic models of hepatocellular carcinoma [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A010.