Research Program
Cell Surface Interactions

Project #
CSI17

Participating Faculty:	C. Fischbach-Teschl and L. Estroff
NBTC Students/Postdocs:	Amy Richter (Estroff)
Other Students/Postdocs:	Debra (Deng) Lin and Siddharth Pathi

Objectives

1) To determine the effect of the nanostructure (i.e., size, shape [aspect ratio]), composition, and crystallinity) of hydroxyapatite nanocrystals on the tumorigenic and osteolytic activity of breast cancer cells using a 3-D polymeric culture platform.

2) To assess whether breast cancer cells regulate the biologic growth of hydroxyapatite nanocrystals in a manner that further promotes osteolytic metastasis using a tumor cell incorporating double-diffusion system.

Methods

1) Nanocrystalline Hydroxyapatite Scaffolds for 3-D cell culture. The Fischbach-Teschl group has developed a microporous poly(lactice-co-glycolide) (PLG) scaffold system to recreate microenvironmental conditions representative of tumors in vivo (Fig. 1). To simulate conditions present in bone (for studying tumor metastasis), we incorporate hydroxyapatite nanocrystals, synthesized by the Estroff group (Fig. 2 (left)). These particles, formed via a hydrothermal reaction, have defined aspect ratios and crystallinities.

2) Cell Culture of Human Breast Cancer Cells. MDA-MB231 cells are used as a model for metastatic breast cancer cells. These cells are cultured within the biomineralized 3-D scaffolds and assayed for characteristics of metastatic cell growth and osteolytic potential (e.g., analysis of up-regulation of osteoclastic IL-8), a typical characteristic of breast cancer bone metastasis (Fig. 2 (right)).

3) Double-Diffusion System and Culture of MDA-MB231 Breast Cancer Cells. To more completely understand the nanoscale dynamics of breast cancer bone metastasis, we are investigating the mechanisms and effects by which metastatic tumor cells influence bone mineral formation. To this end, we examine the effect of tumor cells on the formation of nanocrystalline hydroxyapatite, as well as the effect of ongoing mineralization on the metastatic activity of the tumor cells themselves. Estroff has adapted a dynamic double-diffusion system for studying mineralization under biologically relevant conditions (Fig. 3). A plug of cells is incorporated at the center of the gel tube, which is encased within a gas-permeable silicone tube.

Summary
Breast cancer frequently metastasizes to bone where it leads to osteolysis and poor clinical prognosis; however, the underlying mechanisms and effects remain unclear. We hypothesize that the nanostructure of the bone mineral matrix regulates bone metastasis by controlling breast cancer cell behavior. To address this hypothesis we will integrate scaffold-based human metastasis models, engineered hydroxyapatite nanoparticles of systematically varying physicochemical composition and structure, and a double diffusion system of biomimetic mineralization. This work will elucidate the functional relationships between the nano-scale characteristics of hydroxyapatite, mammary tumor cell behavior, and osteolytic bone metastasis, potentially establishing a new paradigm for the induction and progression of tumor-bone interactions during metastatic breast cancer. Additionally, these studies will be critical to the rationale design of safe biomineralized matrices for bone and tooth regeneration by defining specific nanoscale characteristics of apatite that must be considered for other applications due to their potential tumorigenic properties.

Goals & Accomplishments

• Preparation of biomineralized 3-D culture scaffolds (Fig. 1).
• Preparation of hydroxyapatite nanocrystals with control over aspect ratio and crystallinity (Fig. 2, left).
• Preliminary data suggests that parameters such as crystallinity of crystal aspect ratio affect the secretion of IL-8 by breast cancer cells (Fig. 2, right).