Abstract 3946: Saracatinib inhibits human prostate cancer cell bone metastases in a xenograft model.

Background: In advanced prostate cancer (PCa), cells escape from the primary tumor and enter the bloodstream. These circulating cells preferentially target the axial skeleton and can form bone metastases. This results in weakened bones, spinal compressions, and fractures. Patients experience pain and other bone-related events that increase mortality and decrease quality of life. Therefore, identifying factors that lead to cancer progression, such as areas of hypoxia in the tumor microenvironment, will be crucial to developing strategies to inhibit the spread of malignant cells to distant sites. Efforts to interfere with early steps in the metastatic cascade have led to the development of promising small molecule targeting agents. One such agent is the tyrosine kinase inhibitor (TKI), saracatinib, shown to inhibit activation of Src and other kinases thought to be involved in cancer cell dissemination. The aim of these studies was to use aggressive, human PCa cells (PC-3ML), selected for their ability to form bone tumors in mice, to investigate the effects of various agents shown to interfere with metastatic activity. Methods: PC-3ML cells (a gift from A. Fatatis, Drexel University), expressing Green Fluorescent Protein (GFP) and luciferase (Luc), were injected into the left ventricle of male nude mice, using a closed-chest technique, to ensure that they bypass the lungs and circulate throughout the body. Prior to injection, cells were subjected to either hypoxia for 6 or pretreated for 24 hr with saracatinib. Immediately after injection, mice were placed in an in vivo imaging system (Xenogen IVIS, Caliper Life Sciences) to verify that the cells had disseminated throughout the body. This ensured that only successfully injected mice were randomization to treatment groups. Additional images were collected weekly thereafter to monitor the location and development of tumor nodules. Three weeks post-injection, mice were euthanized and tissues expressing Luc were collected, including bones of the legs, mandible and spine. The bones were fixed and decalcified, followed by flash freezing in OCT freezing medium. Serial frozen sections were then evaluated for GFP expressing cells using a fluorescent stereoscope. Results: Tumor cell deposits were almost always found in bone, frequently in the tibial-femoral joint, scapula-humerus joint, spine, and mandible. Mice injected with PC-3ML cells pretreated with saracatinib had fewer metastases and cells exposed to hypoxia for 6 hr prior to injection had more metastases, on average, than control cells, as predicted by in vitro assays. Conclusions: Intracardiac injection of GFP- and Luc- labeled PC-3ML cells resulted in skeletal metastases that could be monitored in mice over time using an in vivo imaging system and in tissue sections by fluorescent microscopy. The studies verified in vitro data showing that hypoxia can enhance, and saracatinib inhibit, metastatic activity. Citation Format: Lori P. Rice, Christine Pampo, Sharon Lepler, Dietmar W. Siemann. Saracatinib inhibits human prostate cancer cell bone metastases in a xenograft model. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3946. doi:10.1158/1538-7445.AM2013-3946