Chick chorioallantoic membrane (CAM) assays as a model of xenografts derived from circulating cancer stem cells from breast cancer patients.

Abstract

e15596 Background: Circulating cancer stem cells (cCSCs) are a rare fraction of circulating tumor cells with a profile of stemness, resistance to chemotherapy and the capacity to generate metastases. Patient-derived xenografts (PDX) are an increasingly noticed tool in oncology, providing biologically meaningful models of many cancer types, and potential platforms for the development of precision oncology approaches. Commonly, mouse models are used for the in vivo assessment of potential new therapeutic targets in cancers. However, given the high cost, time investment, and legal obligations of such animal models CAM assays are an attractive alternative. Methods: In this study, primary cultures from circulating cancer stem cells were established using sphere-forming assays. Subsequently, tumorspheres were transplanted onto the CAM membrane of fertilized chicken eggs to form secondary microtumors. Results: We have developed an innovative in vitro platform for cultivation of CSCs from peripheral blood of breast cancer patients. The number of tumorspheres increased significantly with tumor progression. Patients with metastatic disease had statistically more tumorspheres as compared to patients without metastasis (30 vs 10/100µl blood, p < 0.05). Patients with multiple metastases had more tumorspheres compared to patients with single metastases (30 vs 60/100µl blood, p < 0.05). The number of tumorspheres was positively correlated with Ki-67, Her2 status and grade score in primary breast tumors. Tumorspheres showed self-renewal, growth potentials, invasion and differentiation in vivo. Their transplantation on CAM membranes resulted in the rapid formation of microtumors in many cases. These tumors pathologically closely resembled the primary tumor. The success rate of PDX was positively correlated with aggressiveness and proliferation capacity of the primary tumor. Conclusions: The number of tumorspheres cultured from peripheral blood of cancer patients and the success rate of establishing PDX directly reflect the aggressiveness and proliferation capacity of the primary tumor. A CAM-based PDX model using circulating cancer stem cells provides a fast, low-cost, easy to handle and powerful preclinical platform for drug screening, therapy optimization, and biomarker discovery.