An important step in the process of metastasis from the primary tumor is invasive spread into the surrounding stroma. Using an in vivo invasion assay, we have previously shown that imposed gradients of epidermal growth factor (EGF) or colony-stimulating factor-1 (CSF-1) can induce invasion through an EGF/CSF-1 paracrine loop between cancer cells and macrophages. We now report that invasion induced by other ligands also relies on this EGF/CSF-1 paracrine invasive loop. Using an in vivo invasion assay, we show that MTLn3 breast cancer cells overexpressing ErbB3 exhibit enhanced invasion compared with control MTLn3 cells in response to the ErbB3 ligand HRG-β1. The invasive response of both MTLn3-ErbB3 and transgenic MMTV-Neu tumors to HRG-β1 is inhibited by blocking EGF receptor, CSF-1 receptor, or macrophage function, indicating that invasiveness to HRG-β1 is dependent on the EGF/CSF-1 paracrine loop. Furthermore, we show that CXCL12 also triggers in vivo invasion of transgenic MMTV-PyMT tumors in an EGF/CSF-1–dependent manner. Although the invasion induced by HRG-β1 or CXCL12 is dependent on the EGF/CSF-1 paracrine loop, invasion induced by EGF is not dependent on HRG-β1 or CXCL12 signaling, showing an asymmetrical relationship between different ligand/receptor systems in driving invasion. Our results identify a stromal/tumor interaction that acts as an engine underlying invasion induced by multiple ligands. [Cancer Res 2009;69(7):3221–7]