Research Article Extracellular calcium promotes the migration of breast cancer cells through the activation of the calcium sensing receptor Zuzana Saidak ⁎, Cedric Boudot, Rachida Abdoune, Laurent Petit, Michel Brazier, Romuald Mentaverri, Said Kamel INSERM ERI-12 EA 4292, University of Picardie Jules Verne, 1 Rue des Louvels, Amiens 80037, France ARTICLEINFORMATION ABSTRACT Article Chronology: Received 19 December 2008 Revised version received 3 March 2009 Accepted 5 March 2009 Available online 13 March 2009 Breast cancer is the most frequent form of cancer in women, with the highest incidence of metastasis to the bone. The reason for the preferential destination to the bone is believed to be due to chemoattractant factors released during bone resorption, which act on the cancer cells facilitating their metastasis. One of the factors released during osteolysis that may mediate breast cancer bone localization is Ca 2+ . Here, we show that extracellular Ca 2+ (Ca 2+ o ) acting via the calcium-sensing receptor (CaSR), greatly promotes the migration of bone-preferring breast cancer cells. In Boyden Chamber and Scratch Wound migration assays, an increase in breast cancer cell migration was observed at 2.5 mM and 5 mM Ca 2+ o compared to basal levels for three of the four breast cancer cell lines tested. However, a significantly greater migratory response was observed for the highly bone metastatic MDA-MB-231 cells, compared to the MCF7 and T47D, which have a lower metastatic potential in vivo. The BT474 cells, which do not metastasize to the bone, did not respond to elevated concentrations of Ca 2+ o in the migration assays. Inhibition of either ERK1/2 MAPK or phospholipase Cβ (PLCβ) led to an abolition of the Ca 2+ o -induced migration, implicating these pathways in the migratory response. Knockdown of the CaSR by siRNA resulted in an inhibition of the Ca 2+ o -induced migration, demonstrating the involvement of this receptor in the effect. These results suggest that the activation of the CaSR by elevated Ca 2+ o concentrations, such as those found near resorbing bone, produces an especially strong chemoattractant effect on bone metastatic breast cancer cells toward the Ca 2+ -rich environment. © 2009 Elsevier Inc. All rights reserved. Keywords: Calcium sensing receptor Breast cancer Extracellular calcium Migration Introduction Breast cancer is the most frequent form of cancer in women [1,2], with a high incidence of metastasis to the bone. Approximately, 75% of patients who develop advanced breast cancer will have secondary tumor in the bone, with the majority being osteolytic [3–5]. Development of bone metastases leads to numerous adverse effects, including severe pain, a high risk of fractures and hypercalcemia, and greatly increased mortality rates [5,6], making the field of bone metastasis a crucial area for further investigation, EXPERIMENTAL CELL RESEARCH 315 (2009) 2072 – 2080 ⁎ Corresponding author. Fax: +33 3 22 82 54 25. E-mail addresses: zuzana.saidak@u-picardie.fr (Z. Saidak), cedric.boudot@u-picardie.fr (C. Boudot), rachida.abdoune@yahoo.fr (R. Abdoune), laurent.petit@u-picardie.fr (L. Petit), michel.brazier@u-picardie.fr (M. Brazier), romuald.mentaverri@u-picardie.fr (R. Mentaverri), said.kamel@u-picardie.fr (S. Kamel). Abbreviations: CaSR, Calcium Sensing Receptor; Ca2+o, extracellular calcium; GPCR, G-protein coupled receptor; PLCβ, phospholipase Cβ; ERK1/2, Extracellular signal-regulated kinase 1/2; PTH, parathyroid hormone; PTHrP, parathyroid hormone-related protein; BSA, bovine serum albumin; PBS, phosphate buffered saline; QPCR, quantitative PCR; siRNA, small interfering RNA; shRNA, small hairpin RNA; RANK, receptor activator for nuclear factor κB; RANKL, receptor activator for nuclear factor κB ligand 0014-4827/$ – see front matter © 2009 Elsevier Inc. All rights reserved. doi:10.1016/j.yexcr.2009.03.003 available at www.sciencedirect.com www.elsevier.com/locate/yexcr