Collective cancer cell invasion in contact with fibroblasts through integrin-α5β1/fibronectin interaction in collagen matrix.

Interaction of cancer cells with cancer-associated fibroblasts (CAFs) plays critical roles in tumor progression. Recently we proposed a new tumor invasion mechanism in which invasive cancer cells individually migrate on elongate protrusions of CAFs (CAF fibers) in 3-D collagen matrix. In this mechanism, cancer cells interact with fibronectin fibrils assembled on CAFs mainly through integrin-α5β1. Here we tested whether this mechanism is applicable to the collective invasion of cancer cells, using two E-cadherin-expressing adenocarcinoma cell lines, DLD-1 (colon) and MCF-7 (breast). When hybrid spheroids of DLD-1 cells with CAFs were embedded into collagen gel, DLD-1 cells collectively but very slowly migrated through the collagen matrix in contact with CAFs. Epidermal growth factor and tumor necrosis factor-α promoted the collective invasion, possibly by reducing the E-cadherin junction, as did the transforming growth factor-β inhibitor SB431542 by stimulating the outgrowth of CAFs. Transforming growth factor-β itself inhibited the cancer cell invasion. Efficient collective invasion of DLD-1 cells required large CAF fibers or their assembly as stable adhesion substrates. Experiments with function-blocking Abs and siRNAs confirmed that DLD-1 cells adhered to fibronectin fibrils on CAFs mainly through integrin-α5β1. Anti-E-cadherin Ab promoted the single cell invasion of DLD-1 cells by dissociating the E-cadherin junction. Although the binding affinity of MCF-7 cells to CAFs was lower than DLD-1, they also collectively invaded the collagen matrix in a similar fashion to DLD-1 cells. Our results suggest that the direct interaction with CAFs, as well as environmental cytokines, contributes to the collective invasion of cancers.