Establishment of a primary cell culture of bone metastasis from urothelial carcinoma.

Abstract

e17030 Background: Urothelial carcinoma (UC) accounts for 90% of cases of bladder cancer. In advanced UC diagnosis, skeletal involvement is frequent. Here we report the establishment of a primary culture of bone metastasis (BM) from High Grade UC derived from 81 years-old patient with a previous diagnosis of papillary urothelial cancer in 2016. After 2 years, patient relapsed to bone and visceral sites and because of poor clinical condition, patient underwent palliative best supportive care. Methods: Patient-derived bone metastatic cell culture was obtained from a surgical specimen. Tumor cells were isolated by collagenase digestion. Cells were seeded on in vitro 2D plates or into 3D collagen scaffolds composed of bovine collagen type I. The establishment of tumor cell isolation was confirmed through the evaluation of cytomorphologic features and positive pan-citokeratin staining. Then, drug treatments was performed and cell survival was evaluated. The study was approved by Ethical Committee and patient signed an informed consent. Results: After a long-term culture, we were able to isolate from a 3D scaffold a tumor clone that successfully growth until passage 20. Stabilized cells generate spheroid-like aggregates, recreating acinar-structures typical of the primary papillary urothelial tumor. Next, we treated cells with: Gemcitabine (Gem), Carboplatin (Carbo), Docetaxel, Carbo+Gem and Bone targeted drugs (Zoledronic Acid and Denosumab). The most effective treatment was Gem+Carbo, in particular 24% of survival in 2D platform and 87% on 3D collagen scaffold. For all treatments, cells cultured on 3D scaffold were more resistant to drugs, mimicking more closely the in-vivo condition. Conclusions: We were able to isolate and establish a BM primary culture from UC using a 3D in-vitro collagen scaffold. This system can recreate microenvironmental conditions more similar to in-vivo ones and it promoted the isolation of tumor cell clones from stromal components of the heterogeneous primary culture. This cell line could be useful to investigate the molecular and genetic profile in order to identify promising molecular targets and to better understard the natural history of BM from UC.