Abstract IA2: Microenvironmental control of bone metastasis
Authors: Provot Sylvain , Brenot Audrey , Casbon Amy-Jo , Yu Ying , Werb Zena 
Published: August 2014

Breast cancer is often associated with a poor prognosis largely due to its tendency to metastasize. Breast cancer preferentially metastasizes to the skeleton. Since Paget's seed and soil hypothesis from more than a century ago, it is known that bones constitute a very hospitable soil that attracts and allows breast cancer cells to thrive. Surprisingly, however, only a few studies have addressed the role of the bone microenvironment in bone metastasis, and most of them emphasized the role of osteoclasts (bone-destructing cells). Osteoclasts are known to stimulate bone metastasis because of their bone resorbing activity, which releases into the bone marrow space growth factors normally embedded in the bone matrix. On the other hand, the role of osteoblasts (bone-making cells) in this process has not been addressed directly. There is growing evidence that osteoblasts' function is not limited to making bone. In particular, these cells regulate various aspects of hematopoiesis, insulin secretion in the pancreas, or even male fertility in mice. We hypothesized that osteoblasts play an important role in breast cancer metastasis to bone. We tested this possibility by inoculating a breast cancer cell line (PyMT-FVB-Luc+ cells) into either the mammary fat pad, or directly into the blood stream of syngeneic recipient mice, which presented either an increased or a decreased osteoblast number. Hypoxia and the hypoxia inducible factor 1alpha (Hif-1alpha) play a fundamental role in bone by stimulating the differentiation of the osteoblasts in hypoxic niches where metastatic cells coming from distant tumors are often found. However, the role of the hypoxic microenvironment in bone metastasis is unknown. We used a conditional knockout approach to suppress either Hif-1alpha or von Hippel Lindau (VHL, an E3 ubiquitin-ligase that targets Hif-1alpha for degradation) specifically in osteoblasts (ΔHif-1alphaOB and ΔVHLOB mice). VHL suppression leads to ectopic expression of Hif-1alpha, and thus functions as a gain-of-function model for Hif-1alpha. Tissue specific ablation of Hif-1alpha and VHL in osteoblasts was performed by mating floxed Hif-1alpha or VHL mice, with mice expressing the Cre recombinase under the control of the osterix promoter, which is solely activated in immature osteoblasts. ΔHif-1alphaOB mice presented a decreased number of osteoblasts and reduced bone mass. Conversely, ΔVHLOB mice exhibited a dramatic increase in osteoblast number and bone mass. Intracardiac injection of PyMT-FVB-Luc+ cells into these loss- or gain-of-function models revealed that osteoblasts directly or indirectly stimulate breast cancer metastasis to bone. Strikingly, our results also show that osteoblasts produce systemic changes, controlling breast cancer metastasis to the lungs, as well as the growth of primary mammary tumors. These results were obtained by either transplanting PyMT-FVB-Luc+ cells into the mammary gland, or by tail vein (intravenous) injections. Thus, osteoblasts can affect distant tissues, well beyond the bone microenvironment. By providing the first evidence that the skeleton exerts a systemic control of breast cancer growth and dissemination, our study expands the biological importance of this organ.

Citation Format: Sylvain Provot, Audrey Brenot, Amy-Jo Casbon, Ying Yu, Zena Werb. Microenvironmental control of bone metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr IA2.