Abstract B21: Establishment of a novel spontaneous prostate cancer liver metastasis model
Authors: Zoggel Hanneke J.A.A. van , Erkens-Schulze Sigrun E. , Ridder Corrina M.A. De , Teubel Wilma , Saleem Hamza , Roobol Stefan J. , Hoeben Sander A.H. , Ridwan Yanto R. , Haeck Joost C. , Weerden Wytske M. van 
Published: March 2017

Abstract
Objective: Prostate cancer (PCa) preferentially spreads to bone with additional metastasis to regional lymph nodes, lung and liver in a subset of patients. Understanding the determining factors of this metastatic pattern and to elucidate the involved pathways and interplay between epithelial and the stromal niche, we established a patient-derived xenograft (PDX) model showing spontaneous metastatic spread to all relevant organs and metastatic lesions in liver.
Methods: PDX of PCa were obtained from NMRI tumor-bearing mice and 3D cell cultures were established by collagenase treatment o/n in prostate growth medium (PGM) containing 2% fetal calf serum. 3D cultures of PDX PC339 were established (PC339C) that could be passaged and stored in liquid nitrogen. PC339C was transfected with a lentiviral construct containing luc2-GFP fused protein (M21) and upon FACS-sorting the PC339C-M21 was retrieved. PC339C-M21 cells were inoculated subcutaneously in NSG mice and when tumors were established mice underwent tumorectomy (700-1000 mm3) to extend the life span of the animal and allow metastastic outgrowth. Mice were monitored weekly for metastatic outgrowth using IVIS, microCT and MRI. At sacrifice, M21-positive organs, including liver, lymph nodes, lung and bone, were sampled. Part of the tissue was used for culturing, part was fresh frozen for RNA and protein isolation to allow qPCR and western blot analyses, and part was formalin-fixed and paraffin-embedded for histology and immunohistochemistry.
Results: PC339C-M21 tumors were established in NSG mice and after tumorectomy a spontaneous liver metastasis was established. Culturing of this tumor resulted in the PC339C-M21-L cell line. This cell line was inoculated subcutaneously in NSG mice to result in tumor formation and spontaneous liver metastasis in all mice. Tumorectomy was performed at 25-30 days after inoculation. Luciferase-positive signals in liver were detected by IVIS as early as 7 days after tumorectomy with MRI showing first visible lesions at 21 days and microCT at 28 days. 3D reconstructed microCT images and MRI-detected lesions confirmed the signals and location as seen by IVIS. When inoculated orthotopically, liver metastatic lesions became visible by IVIS at 40 days, which was not different from the subcutaneous tumors. However, these mice could not be followed longer due to the orthotopic tumor burden. On top of the PC339C-M21-L cell line, culturing of luciferase-positive organs resulted in cell lines from lymph node (PC339C-M21-L-LN), lung (PC339C-M21-L-LU), and bone (PC339C-M21-L-B) together constituting a cell line panel of metastatic PCa. This cell line panel is being further characterized in vitro and in vivo for their unique properties and metastatic potential.
Conclusion: A new spontaneous PCa liver metastasis xenograft model, PCL339-M21-L, was generated. This model was extended to yield a panel of metastatic sublines from different organ origin. Together, this platform may serve as a tool to study the biology of spontaneous metastatic outgrowth in liver and other organs. Furthermore, we demonstrated the feasibility of non-invasive imaging techniques to follow metastatic spread and outgrowth allowing to monitor responses to anti-cancer therapies.
Citation Format: Hanneke J.A.A. van Zoggel, Sigrun E. Erkens-Schulze, Corrina M.A. De Ridder, Wilma Teubel, Hamza Saleem, Stefan J. Roobol, Sander A.H. Hoeben, Yanto R. Ridwan, Joost C. Haeck, Wytske M. van Weerden{Authors}. Establishment of a novel spontaneous prostate cancer liver metastasis model. [abstract]. In: Proceedings of the AACR Special Conference: Patient-Derived Cancer Models: Present and Future Applications from Basic Science to the Clinic; Feb 11-14, 2016; New Orleans, LA. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(16_Suppl):Abstract nr B21.