F06 - Integrin a5 is an independent prognosis factor and a potential therapeutic target for breast cancer bone metastasis

Abstract

Background: Using an extensive bioinformatic approach we identified integrin &agr;5 subunit as a novel potential target to treat bone dissemination from breast cancer. Aim of this study is to confirm the value of this target Material and methods: Integrin &agr;5 mRNA expression was quantified by qRT-PCR in 427 breast cancer patients. &agr;5 protein levels on primary tumor was correlated with presence of disseminated tumor cells (DTCs) in bone marrow at the time of surgery in further 268 breast cancer patients. BALB/c mice were injected with human MDA-MB-231 &agr;5 silenced or scramble. Animals were killed on day 14 after tumor cell inoculation; lung and bone marrow were flushed from the hind limbs for DTC colony assay. Alternatively mice were treated with an anti a5 moAb (M200) or vehicle. Treatment was performed 3 times per week starting the day before intra-arterial inoculation with luciferase expressing human MDA-MB-231/B02 breast cancer cells, which selectively metastasize to bone. In an additional protocol, animals were culled on day 7 after tumor cell inoculation, and the bone marrow was flushed for DTC colony assay. Results: An high a5 expression was associated with shorter bone metastasis-free survival, both in univariate (P = 0.024) and multivariate analysis (P = 0.04). Moreover we found significant (P = 0.039) positive association between a5 protein expression on primary cancer and presence of DTCs in bone marrow. Taking advantage of the ability of the MDA-MB-231 cancer cell to metastatize both in bone and lung, we found that abrogating a5 gene function dramatically reduced the number of bone micrometastases (P = 0.015) without affecting lung dissemination. In addition treatment of MDA-MB-231/B02 injected animals with M200 antibody significantly delayed the onset of skeletal lesions (P = 0.02) and caused a 50% reduction in the extent of osteolytic lesions (P = 0.038), compared to vehicle. This difference was accompanied with a sharp reduction of tumor burden (P = 0.02), as determined by bioluminescence. Histomorphometric analysis of metastatic legs showed that M200 treatment decreased skeletal tumor burden (P = 0.027) and increased the bone volume (P = 0.02), compared to vehicle. Additionally, the number of DTC colonies from M200-treated mice was dramatically decreased compared with vehicle (P< 0.001). Conclusions: Our results suggest that &agr;5 integrin expression in breast cancer cells facilitates bone marrow micrometastasis formation and the subsequent development of osteolytic lesions.