Hematogenous metastasis requires tumor cells to detach from primary tumor into blood/lymphatic circulation and extravasate. Tumor cells in the blood circulation system, named circulating tumor cells (CTCs), are in a suspension state, with unique cytoskeletal structure and molecular phenotype different from primary tumor cells. The aim of this study is to assess the impact of suspension state on the metastatic potential of breast cancer cells (BCCs) and study its underlying mechanism. Methods: BCCs were cultured on low-adhesion plates to mimic the suspension state. Conventional adherent culture BCCs were used as the control. This study examined the metastatic potential of adherent and suspension BCCs in vitro and in vivo. RNA sequencing analysis, siRNA, and inhibitors were used to determine the underlying molecular mechanism. Results: The suspension state significantly increased the metastatic potential of BCCs, but slightly suppressed their tumor growth. RNA sequencing analysis revealed that the suspension state resulted in an acquisition of unique molecular signature enriched in pro-metastatic and tumor-suppressive genes. Specifically, prostaglandin-endoperoxide synthase 2 (PTGS2), which encodes protein cyclooxygenase-2 (COX-2), was identified as a highly up-regulated gene in suspension state compared with adherent cultured BCCs. Inhibition of the catalytic activity of COX-2 by celecoxib markedly suppressed suspension-increased migration and invasion of BCCs. In addition, knock-down of COX-2 by siRNA reduced the experimental lung metastasis formation of suspension cultured BCCs, which was associated with a remarkable decrease in retention and survival of BCCs in lungs of mice in the early stage of metastasis. Activation of Ca2+/calcineurin (CaN)/nuclear factor of activated T cells (NFAT) pathway and disruption of cytoskeleton contributed to the COX-2 up-expression by suspension state. Conclusions: Our results demonstrate that suspension state plays an important role in the metastatic potential of CTCs, and suggest a potential application of COX-2 inhibitor for anti-metastasis.