Tumor-α9β1 integrin-mediated signaling induces breast cancer growth and lymphatic metastasis via the recruitment of cancer-associated fibroblasts

Tumor-derived matricellular proteins such as osteopontin (OPN) and tenascin-C (TN-C) have been implicated in tumor growth and metastasis. However, the molecular basis of how these proteins contribute to tumor progression remains to be elucidated. Importantly, these matricellular proteins are known to interact with alpha 9 beta 1 integrin. Therefore, we hypothesized that tumor-derived alpha 9 beta 1 integrin may contribute to tumor progression. To clarify the roles of alpha 9 beta 1 integrin in tumor growth and lymphatic metastasis, we used an inhibitory anti-human alpha 9 beta 1 integrin antibody (anti-h alpha 9 beta 1 antibody) and a alpha 9 beta 1 integrin-positive human breast cancer cell line, MDA-MB-231 luc-D3H2LN (D3H2LN), in vitro functional assays, and an in vivo orthotopic xenotransplantation model. In this study, we demonstrated that tumor, but not host alpha 9 beta 1 integrin, contributes to tumor growth, lymphatic metastasis, recruitment of cancer-associated fibroblasts (CAFs), and host-derived OPN production. We also found that CAFs contributed to tumor growth, lymphatic metastasis, and host-derived OPN levels. Consistent with those findings, tumor volume was well-correlated with numbers of CAFs and levels of host-derived OPN. Furthermore, it was shown that the inoculation of D3H2LN cells into mammary fat pads with mouse embryonic fibroblasts (MEFs), obtained from wild type, but not OPN knock-out mice, resulted in enhancement of tumor growth, thus indicating that CAF-derived OPN enhanced tumor growth. These results suggested that tumor alpha 9 beta 1-mediated signaling plays a pivotal role in generating unique primary tumor tissue microenvironments, which favor lymphatic metastasis and tumor growth.