G. Oshima1,2, S. C. Wightman1, A. Uppal1, J. Oskvarek2, M. Stack1, X. Huang2, T. E. Darga2, M. C. Posner1, N. Khodarev2, R. R. Weichselbaum2 1University Of Chicago,Department Of Surgery,Chicago, IL, USA 2University Of Chicago,Department Of Radiation & Cellular Oncology, Ludwig Center For Metastasis Research,Chicago, IL, USA
Introduction: Patients with colorectal cancer (CRC) often present with liver metastases, which, despite systemic treatment, frequently results in a fatal outcome. Between 5 to 20% of patients with limited numbers of hepatic metastases and slow rates of progression/recurrence (oligometastases) are successfully treated with local treatment approaches, with or without systemic therapy. Yet, little is known about molecular basis of oligometastatic disease. Animal models are critical to study pathophysiological mechanisms and new treatments for patients with metastatic CRC. We propose here a xenogenic animal model of hepatic metastasis of CRC that simulate oligo- and polymetastases,
Methods: We generated a panel of monoclonal HCT116-derived cell lines double-labeled with luciferase and tdTomato protein using lentiviral-based gene delivery. Hepatic tumors were generated by injecting 2 million cells into spleens in mice. Five minutes post-injection splenectomies were performed to avoid carcinomatous peritonitis and residual growth in spleen. Bioluminescent signals were measured weekly using the Xenogen IVIS 200 (MA, United States) imaging system. Two to 4 weeks after injections, livers were harvested and ex vivo fluorescent intensities of liver tumors were quantified. The number and size of liver tumors were directly measured to see the correlation between fluorescent intensities and macroscopic disease. Twenty individual monoclonal cell lines were tested to select candidates for oligo- and polymetastatic phenotype. Diffuse Luminescent Imaging Tomography (DLIT) was performed for evaluation of tumor burden and organ distribution using real-time 3D reconstruction of bioluminescence signals.
Results: Monoclonal cell lines varied in the ability to form liver metastases with 2 cell lines demonstrating oligometastatic phenotype (O1 and O2) and 2 other cell lines-polymetastatic phenotype (P1 and P2). Bioluminescent intensities of P1 and P2 were higher than O1 and O2 at 1 to 3 weeks after injection. Mean tumor number of P1 was higher than P2, O1 and O2 (p < 0.05). Mean tumor size of P2 was larger than P1, O1 and O2 (p < 0.05). Tumor burden estimated by tumor number and size correlated with bioluminescent intensity and fluorescent intensity (R = 0.99). DLIT showed tumor distribution and size dependent signal intensity comparable to macroscopic findings.
Conclusion: An animal model of metastatic CRC to the liver can reproducibly mimic oligo- and polymetastatic CRC in humans. Cell lines double-labeled with luciferase and tdTomato protein can be used as a reliable method to evaluate tumor burden using live imaging which enable sequential monitoring in the same mice and detailed information about tumor size and distribution in secondary site(s). This animal model can be used to improve pre-clinical testing of novel therapies. It may also facilitate evaluation of new biological mechanisms discriminating oligo-and polymetastatic disease in liver.