Abstract 437: Mena at the nexus of chemotaxis and haptotaxis during tumor progression
Authors: Oudin Madeleine J. , Jonas Oliver , Kosciuk Tatiana , Broye Liliane C. , Wyckoff Jeff , Miller Miles A. , Lussiez Alisha , Asokan Sreeja , Langer Robert , Lauffenburger Douglas , Bear James E. , Gertler Frank B. 
Published: August 2015

Abstract
The most well studied mode of cell motility for metastasizing tumor cells is chemotaxis, the directional movement of cells attracted to a source of soluble cues. In contrast, haptotaxis, migration of cells on gradients of substrate-bound factors, remains poorly understood. Due to the high expression of extracellular matrix (ECM) proteins such as collagen and fibronectin (FN) in metastatic tumors, it is possible that haptotaxis along ECM gradients could also be driving invasion. Mena, an actin regulatory protein, plays an important role in cell motility and is upregulated in various cancers, where Mena, and in particular the MenaINV isoform, potentiates chemotactic and invasive responses to EGF. All Mena isoforms bind directly to the α5 subunit of the α5β1 integrin, a FN receptor. We hypothesized that the pro-metastatic effect of MenaINV arises from increases in tumor cell responses to specific chemotactic and haptotactic cues, including EGF and FN.
To study haptotaxis, MDA-MB231 breast adenocarcinoma cells were plated on gradients of fluorescently-labeled FN in 2D or in 3D collagen gels, generated within a PDMS microfluidic device. Wild type cells expressing low levels of endogenous Mena failed to respond to a FN gradient, but migrated equally well in all directions irrespective of the FN gradient. Surprisingly, ectopic expression of Mena in these cells conferred the ability to respond to FN gradients, with cells exhibiting haptotactic response towards higher concentrations of FN in 2D and 3D. Interestingly, cells expressing MenaINV were able to haptotax towards FN concentrations 4 times higher then cells expressing Mena, while reorganizing and recruiting both collagen and FN surrounding them. MenaINV-dependent haptotaxis was ablated by deletion of the α5 binding site within Mena or treatment with an α5 function blocking antibody, but was unaffected by inhibition of another FN receptor, αVβ3.
Using intravital imaging of mouse mammary tumors, we visualized haptotaxis of MenaINV-expressing cells towards gradients of fluorescently labelled FN released locally from implanted microsccale devices. Tumor cells expressing MenaINV lacking the α5 binding site did not haptotax, displayed decreased motility and metastasis to the lung. This was also accompanied by a striking change in the organization of the ECM surrounding the tumor. While investigating the mechanism of haptotaxis, we found that MenaINV-driven invasive responses to FN were also dependent on EGFR signalling and mediated by the phosphatase PTP1B. Finally, using a combination of in vitro and in vivo invasion assays, we show that MenaINV expression promoted synergistic invasive responses to combinations of EGF and FN and also caused increased phosphorylation of Paxilin and FAK at focal adhesions.
Together, our data demonstrate that MenaINV is a shared component of the machinery that mediates both haptotactic responses to FN and chemotactic responses to EGF, two pathways that promote tumor progression.
Citation Format: Madeleine J. Oudin, Oliver Jonas, Tatiana Kosciuk, Liliane C. Broye, Jeff Wyckoff, Miles A. Miller, Alisha Lussiez, Sreeja Asokan, Robert Langer, Douglas Lauffenburger, James E. Bear, Frank B. Gertler. Mena at the nexus of chemotaxis and haptotaxis during tumor progression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 437. doi:10.1158/1538-7445.AM2015-437