Stat3 regulates ErbB-2 expression and co-opts ErbB-2 nuclear function to induce miR-21 expression, PDCD4 downregulation and breast cancer metastasis.

Membrane overexpression of the receptor tyrosine kinase ErbB-2 (MErbB-2) accounts for a clinically aggressive breast cancer (BC) subtype (ErbB-2-positive) with increased incidence of metastases. We and others demonstrated that nuclear ErbB-2 (NErbB-2) also plays a key role in BC and is a poor prognostic factor in ErbB-2-positive tumors. The signal transducer and activator of transcription 3 (Stat3), another player in BC, has been recognized as a downstream mediator of MErbB-2 action in BC metastasis. Here, we revealed an unanticipated novel direction of the ErbB-2 and Stat3 interaction underlying BC metastasis. We found that Stat3 binds to its response elements (GAS) at the ErbB-2 promoter to upregulate ErbB-2 transcription in metastatic, ErbB-2-positive BC. We validated these results in several BC subtypes displaying metastatic and non-metastatic ability, highlighting Stat3 general role as upstream regulator of ErbB-2 expression in BC. Moreover, we showed that Stat3 co-opts NErbB-2 function by recruiting ErbB-2 as its coactivator at the GAS sites in the promoter of microRNA-21 (miR-21), a metastasis-promoting microRNA (miRNA). Using an ErbB-2 nuclear localization domain mutant and a constitutively activated ErbB-2 variant, we found that NErbB-2 role as a Stat3 coactivator and also its direct role as transcription factor upregulate miR-21 in BC. This reveals a novel function of NErbB-2 as a regulator of miRNAs expression. Increased levels of miR-21, in turn, downregulate the expression of the metastasis-suppressor protein programmed cell death 4 (PDCD4), a validated miR-21 target. Using an in vivo model of metastatic ErbB-2-postive BC, in which we silenced Stat3 and reconstituted ErbB-2 or miR-21 expression, we showed that both are downstream mediators of Stat3-driven metastasis. Supporting the clinical relevance of our results, we found an inverse correlation between ErbB-2/Stat3 nuclear co-expression and PDCD4 expression in ErbB-2-positive primary invasive BCs. Our findings identify Stat3 and NErbB-2 as novel therapeutic targets to inhibit ErbB-2-positive BC metastasis.

Soluble factors from the target organ enhance tumor cell angiogenesis: role of laminin SIKVAV sequence.

The ability of tumor cells to respond to microenvironmental factors present in the target organ determines in part the successful development of a metastasis. In a previous work it was demonstrated that the conditioned medium (CM) from lungs of normal mice stimulates in vitro migration, proliferation and uPA activity of cells from a murine mammary adenocarcinoma moderately metastatic to lung. This CM also enhanced local and metastatic tumor growth. Here, we show that lung CM enhanced neovascularization when inoculated together with LM3 tumor cells into the skin of syngeneic mice. A similar tumor-induced angiogenesis response was obtained when lung CM was injected systemically. Western blot analysis of lung CM revealed the presence of some laminin fragments containing the sequence SIKVAV. To determine whether those molecules were responsible for the observed angiogenic effects, the CM was depleted of the peptides containing the SIKVAV sequence. We observed that the SIKVAV-depleted lung CM lost its ability to induce an enhancement of the tumor neovascular response. Our results suggest a role for the target organ in facilitating the neovascularization of tumor cells, probably through the participation of active peptides derived from the proteolytic degradation of the basement membrane component laminin.