Abstract 3206: Three-dimensional hyaluronan hydrogel culture systems facilitate study of the complex microenvironment of bone metastatic prostate cancer cells

The bone marrow microenvironment is a complex milieu, containing various stromal and endothelial cell populations as well as a unique extracellular matrix. To better understand cellular interactions that occur in potentially lethal stage IV prostate cancer (PCa), it is helpful to recreate the bone marrow microenvironment. Traditional two dimensional (2D) cell culture systems do not accurately portray this environment, since culture on plastic activates atypical signaling pathways and makes co-culture of various cell types difficult. To overcome these limitations, we developed novel methods of three-dimensional (3D) culture to study the biology of bone metastatic PCa cells. These 3D culture systems utilize ECM-modified hyaluronan (HA) hydrogels to culture PCa cells and bone marrow stromal cells (BMSC) in a variety of well controlled configurations. The HA hydrogel reflects the low modulus environment of the bone marrow as well as HA's prevalence within the tissue. Various types of PCa cells, including cells derived from patient bone metastases, were cultured in HA hydrogels and viability was maintained as assessed by live/dead assays over time. Because HA is a bio-active signaling molecule that can interact with PCa cells through its receptors, cluster of differentiation 44 (CD44) or receptor for hyaluronan mediated motility (RHAMM), changes in morphology and signaling pathway activation were observed in response to the HA hydrogel culture. To investigate how PCa and BMSC interact, we encapsulated BMSC (HS27a) within a collagen-modified HA hydrogel. These cells formed a reticular network in the 3D matrix, reminiscent of the architecture of bone marrow stroma. When co-cultured with BMSC, bone metastatic PCa cells adhered to the reticular network of stromal cells and exhibited phenotypic changes suggesting neuroendocrine differentiation. We are developing a bilayered HA-based system to model, in 3D, the interface between PCa and the bone marrow stroma. Our model system includes 6 mm diameter disks of HA composed of two well-defined layers, one encapsulating BMSCs and the other containing PCa cells; each cell line can be labeled with a different fluorescent protein. These bilayered disks can be imaged to study the means of invasion of PCa into bone marrow stroma mimicking metastasis. Changes in cell-cell interactions and in cell morphologies in 3D can be imaged without disturbing the dynamic system, allowing for experiments to be monitored over time. These str