Control of SRC molecular dynamics encodes distinct cytoskeletal responses by specifying signaling pathway usage.

Upon activation by different transmembrane receptors, the same signaling protein can induce distinct cellular responses. A way to decipher the mechanisms of such pleiotropic signaling activity is to directly manipulate the decision-making activity that supports the selection between distinct cellular responses. We developed an optogenetic probe (optoSRC) to control SRC signaling, an example of a pleiotropic signaling node, and we demonstrated its ability to generate different acto-adhesive structures (lamellipodia or invadosomes) upon distinct spatio-temporal control of SRC kinase activity. The occurrence of each acto-adhesive structure was simply dictated by the dynamics of optoSRC nanoclusters in adhesive sites, which were dependent on the SH3 and Unique domains of the protein. The different decision-making events regulated by optoSRC dynamics induced distinct downstream signaling pathways, which we characterized using time-resolved proteomic and network analyses. Collectively, by manipulating the molecular mobility of SRC kinase activity, these experiments reveal the pleiotropy-encoding mechanism of SRC signaling.

CCM2-deficient endothelial cells undergo a ROCK-dependent reprogramming into senescence-associated secretory phenotype.

Cerebral cavernous malformation (CCM) is a cerebrovascular disease in which stacks of dilated haemorrhagic capillaries form focally in the brain. Whether and how defective mechanotransduction, cellular mosaicism and inflammation interplay to sustain the progression of CCM disease is unknown. Here, we reveal that CCM1- and CCM2-silenced endothelial cells expanded in vitro enter into senescence-associated secretory phenotype (SASP) that they use to invade the extracellular matrix and attract surrounding wild-type endothelial and immune cells. Further, we demonstrate that this SASP is driven by the cytoskeletal, molecular and transcriptomic disorders provoked by ROCK dysfunctions. By this, we propose that CCM2 and ROCK could be parts of a scaffold controlling senescence, bringing new insights into the emerging field of the control of ageing by cellular mechanics. These in vitro findings reconcile the known dysregulated traits of CCM2-deficient endothelial cells into a unique endothelial fate. Based on these in vitro results, we propose that a SASP could link the increased ROCK-dependent cell contractility in CCM2-deficient endothelial cells with microenvironment remodelling and long-range chemo-attraction of endothelial and immune cells.

Piezo2 channel regulates RhoA and actin cytoskeleton to promote cell mechanobiological responses.

Actin polymerization and assembly into stress fibers (SFs) is central to many cellular processes. However, how SFs form in response to the mechanical interaction of cells with their environment is not fully understood. Here we have identified Piezo2 mechanosensitive cationic channel as a transducer of environmental physical cues into mechanobiological responses. Piezo2 is needed by brain metastatic cells from breast cancer (MDA-MB-231-BrM2) to probe their physical environment as they anchor and pull on their surroundings or when confronted with confined migration through narrow pores. Piezo2-mediated Ca2+ influx activates RhoA to control the formation and orientation of SFs and focal adhesions (FAs). A possible mechanism for the Piezo2-mediated activation of RhoA involves the recruitment of the Fyn kinase to the cell leading edge as well as calpain activation. Knockdown of Piezo2 in BrM2 cells alters SFs, FAs, and nuclear translocation of YAP; a phenotype rescued by overexpression of dominant-positive RhoA or its downstream effector, mDia1. Consequently, hallmarks of cancer invasion and metastasis related to RhoA, actin cytoskeleton, and/or force transmission, such as migration, extracellular matrix degradation, and Serpin B2 secretion, were reduced in cells lacking Piezo2.

The CCM1-CCM2 complex controls complementary functions of ROCK1 and ROCK2 that are required for endothelial integrity.

Endothelial integrity relies on a mechanical crosstalk between intercellular and cell-matrix interactions. This crosstalk is compromised in hemorrhagic vascular lesions of patients carrying loss-of-function mutations in cerebral cavernous malformation (CCM) genes. RhoA/ROCK-dependent cytoskeletal remodeling is central to the disease, as it causes unbalanced cell adhesion towards increased cell-extracellular matrix adhesions and destabilized cell-cell junctions. This study reveals that CCM proteins directly orchestrate ROCK1 and ROCK2 complementary roles on the mechanics of the endothelium. CCM proteins act as a scaffold, promoting ROCK2 interactions with VE-cadherin and limiting ROCK1 kinase activity. Loss of CCM1 (also known as KRIT1) produces excessive ROCK1-dependent actin stress fibers and destabilizes intercellular junctions. Silencing of ROCK1 but not ROCK2 restores the adhesive and mechanical homeostasis of CCM1 and CCM2-depleted endothelial monolayers, and rescues the cardiovascular defects of ccm1 mutant zebrafish embryos. Conversely, knocking down Rock2 but not Rock1 in wild-type zebrafish embryos generates defects reminiscent of the ccm1 mutant phenotypes. Our study uncovers the role of the CCM1-CCM2 complex in controlling ROCK1 and ROCK2 to preserve endothelial integrity and drive heart morphogenesis. Moreover, it solely identifies the ROCK1 isoform as a potential therapeutic target for the CCM disease.

How cells respond to environmental cues - insights from bio-functionalized substrates.

Biomimetic materials have long been the (he)art of bioengineering. They usually aim at mimicking in vivo conditions to allow in vitro culture, differentiation and expansion of cells. The past decade has witnessed a considerable amount of progress in soft lithography, bio-inspired micro-fabrication and biochemistry, allowing the design of sophisticated and physiologically relevant micro- and nano-environments. These systems now provide an exquisite toolbox with which we can control a large set of physicochemical environmental parameters that determine cell behavior. Bio-functionalized surfaces have evolved from simple protein-coated solid surfaces or cellular extracts into nano-textured 3D surfaces with controlled rheological and topographical properties. The mechanobiological molecular processes by which cells interact and sense their environment can now be unambiguously understood down to the single-molecule level. This Commentary highlights recent successful examples where bio-functionalized substrates have contributed in raising and answering new questions in the area of extracellular matrix sensing by cells, cell-cell adhesion and cell migration. The use, the availability, the impact and the challenges of such approaches in the field of biology are discussed.

ICAP-1 monoubiquitylation coordinates matrix density and rigidity sensing for cell migration through ROCK2-MRCKα balance.

Cell migration is a complex process requiring density and rigidity sensing of the microenvironment to adapt cell migratory speed through focal adhesion and actin cytoskeleton regulation. ICAP-1 (also known as ITGB1BP1), a ß1 integrin partner, is essential for ensuring integrin activation cycle and focal adhesion formation. We show that ICAP-1 is monoubiquitylated by Smurf1, preventing ICAP-1 binding to ß1 integrin. The non-ubiquitylatable form of ICAP-1 modifies ß1 integrin focal adhesion organization and interferes with fibronectin density sensing. ICAP-1 is also required for adapting cell migration in response to substrate stiffness in a ß1-integrin-independent manner. ICAP-1 monoubiquitylation regulates rigidity sensing by increasing MRCKα (also known as CDC42BPA)-dependent cell contractility through myosin phosphorylation independently of substrate rigidity. We provide evidence that ICAP-1 monoubiquitylation helps in switching from ROCK2-mediated to MRCKα-mediated cell contractility. ICAP-1 monoubiquitylation serves as a molecular switch to coordinate extracellular matrix density and rigidity sensing thus acting as a crucial modulator of cell migration and mechanosensing.

αvß3 integrins negatively regulate cellular forces by phosphorylation of its distal NPXY site.

BACKGROUND INFORMATION: Integrins are key receptors that allow cells to sense and respond to their mechanical environment. Although they bind the same ligand, ß1 and ß3 integrins have distinct and cooperative roles in mechanotransduction. RESULTS: Using traction force microscopy on unconstrained cells, we show that deleting ß3 causes traction forces to increase, whereas the deletion of ß1 integrin results in a strong decrease of contractile forces. Consistently, loss of ß3 integrin also induces an increase in ß1 integrin activation. Using a genetic approach, we identified the phosphorylation of the distal NPXY domain as an essential process for ß3 integrin to be able to modulate traction forces. Loss of ß3 integrins also impacted cell shape and the spatial distribution of traction forces, by causing forces to be generated closer to the cell edge, and the cell shape. CONCLUSIONS: Our results emphasize the role of ß3 integrin in spatial distribution of cellular forces. We speculate that, by modulating its affinity with kindlin, ß3 integrins may be able to locate near the cell edge where it can control ß1 integrin activation and clustering. SIGNIFICANCE: Tensional homeostasis at the single cell level is performed by the ability of ß3 adhesions to negatively regulate the activation degree and spatial localization of ß1 integrins. By combining genetic approaches and new tools to analyze traction distribution and cell morphology on a population of cells we were able to identify the molecular partners involved in cellular forces regulation.

The RNA-binding protein Mex3b regulates the spatial organization of the Rap1 pathway.

The four related mammalian MEX-3 RNA-binding proteins are evolutionarily conserved molecules for which the in vivo functions have not yet been fully characterized. Here, we report that male mice deficient for the gene encoding Mex3b are subfertile. Seminiferous tubules of Mex3b-deficient mice are obstructed as a consequence of the disrupted phagocytic capacity of somatic Sertoli cells. In addition, both the formation and the integrity of the blood-testis barrier are compromised owing to mislocalization of N-cadherin and connexin 43 at the surface of Sertoli cells. We further establish that Mex3b acts to regulate the cortical level of activated Rap1, a small G protein controlling phagocytosis and cell-cell interaction, through the activation and transport of Rap1GAP. The active form of Rap1 (Rap1-GTP) is abnormally increased at the membrane cortex and chemically restoring Rap1-GTP to physiological levels rescues the phagocytic and adhesion abilities of Sertoli cells. Overall, these findings implicate Mex3b in the spatial organization of the Rap1 pathway that orchestrates Sertoli cell functions.

Substrate Stiffness Combined with Hepatocyte Growth Factor Modulates Endothelial Cell Behavior.

Endothelial cells (ECs) play a crucial role in regulating various physiological and pathological processes. The behavior of ECs is modulated by physical (e.g., substrate stiffness) and biochemical cues (e.g., growth factors). However, the synergistic influence of these cues on EC behavior has rarely been investigated. In this study, we constructed poly(l-lysine)/hyaluronan (PLL/HA) multilayer films with different stiffness and exposed ECs to these substrates with and without hepatocyte growth factor (HGF)-supplemented culture medium. We demonstrated that EC adhesion, migration, and proliferation were positively correlated with substrate stiffness and that these behaviors were further promoted by HGF. Interestingly, ECs on the lower stiffness substrates showed stronger responses to HGF in terms of migration and proliferation, suggesting that HGF can profoundly influence stiffness-dependent EC behavior correlated with EC growth. After the formation of an EC monolayer, EC behaviors correlated with endothelial function were evaluated by characterizing monolayer integrity, nitric oxide production, and gene expression of endothelial nitric oxide synthase. For the first time, we demonstrated that endothelial function displayed a negative correlation with substrate stiffness. Although HGF improved endothelial function, HGF was not able to change the stiffness-dependent manner of endothelial functions. Taken together, this study provides insights into the synergetic influence of physical and biochemical cues on EC behavior and offers great potential in the development of optimized biomaterials for EC-based regenerative medicine.

Microenvironment, tumor cell plasticity, and cancer.

PURPOSE OF REVIEW: Much effort has been devoted to determining how cellular and noncellular components of the tumoral niche initiate and promote cancer development. Cancer cells perceive biochemical signals from components of the extracellular matrix (ECM) and sense physical features, such as matrix stiffness and cell confinement. The past decade has seen a better understanding of the biophysics and mechanobiology associated with cancer cells. Indeed, loss of mechanisms controlling the production, the degradation, and the remodeling of ECM contributes to tumor growth or cell dissemination by affecting cell contractility in response to ECM stiffness and by stimulating mechanical dependence of growth factor activation. RESULTS: Cell plasticity allows adaptative strategies for cancer cells to survive or eventually escape from tumoral environment through modification of the microenvironment-cell interface, internal tension increase, and nuclear deformation partly leading to intratumoral heterogeneity. However, although alteration of the biomechanical properties of the ECM are sufficient to promote cell migration and invasion in cancer cells, this microenvironment can also provide a hospitable niche for tumor dormancy and resistance to cancer therapy. CONCLUSION: The review will focus on how physicochemical properties of ECM might promote tumor growth or cell dissemination or on the contrary maintain quiescent state of cancer cells. It is crucial to clarify the molecular basis of mechanotransduction in the development and progression of tumors to identify new potential biomarkers and anticancer therapeutic targets.

Calcium and calmodulin-dependent serine/threonine protein kinase type II (CaMKII)-mediated intramolecular opening of integrin cytoplasmic domain-associated protein-1 (ICAP-1α) negatively regulates ß1 integrins.

Focal adhesion turnover during cell migration is an integrated cyclic process requiring tight regulation of integrin function. Interaction of integrin with its ligand depends on its activation state, which is regulated by the direct recruitment of proteins onto the ß integrin chain cytoplasmic domain. We previously reported that ICAP-1α, a specific cytoplasmic partner of ß1A integrins, limits both talin and kindlin interaction with ß1 integrin, thereby restraining focal adhesion assembly. Here we provide evidence that the calcium and calmodulin-dependent serine/threonine protein kinase type II (CaMKII) is an important regulator of ICAP-1α for controlling focal adhesion dynamics. CaMKII directly phosphorylates ICAP-1α and disrupts an intramolecular interaction between the N- and the C-terminal domains of ICAP-1α, unmasking the PTB domain, thereby permitting ICAP-1α binding onto the ß1 integrin tail. ICAP-1α direct interaction with the ß1 integrin tail and the modulation of ß1 integrin affinity state are required for down-regulating focal adhesion assembly. Our results point to a molecular mechanism for the phosphorylation-dependent control of ICAP-1α function by CaMKII, allowing the dynamic control of ß1 integrin activation and cell adhesion.

Extracellular matrix rigidity controls podosome induction in microvascular endothelial cells.

BACKGROUND INFORMATION: Podosomes are actin-based structures involved in cell adhesion, migration, invasion and extracellular matrix degradation. They have been described in large vessel endothelial cells, but nothing is known concerning microvascular endothelial cells. Here, we focussed on liver sinusoidal endothelial cells (LSECs), fenestrated microvascular cells that play major roles in liver physiology. Liver fibrosis induces a dedifferentiation of LSECs leading notably to a loss of fenestrae. Because liver fibrosis is associated with increased matrix stiffness, and because substrate stiffness is known to regulate the actin cytoskeleton, we investigated the impact of matrix rigidity on podosome structures in LSECs. RESULTS: Using primary LSECs, we demonstrated that microvascular endothelial cells are able to form constitutive podosomes. Podosome presence in LSECs was independent of cytokines such as transforming growth factor-ß or vascular endothelial growth factor, but could be modulated by matrix stiffness. As expected, LSECs lost their differentiated phenotype during cell culture, which was paralleled by a loss of podosomes. LSECs however retained the capacity to form active podosomes following detachment/reseeding or actin-destabilising drug treatments. Finally, constitutive podosomes were also found in primary microvascular endothelial cells from other organs. CONCLUSIONS: Our results show that microvascular endothelial cells are able to form podosomes without specific stimulation. Our data suggest that the major determinant of podosome induction in these cells is substrate rigidity.

Force tuning through regulation of clathrin-dependent integrin endocytosis.

Integrin endocytosis is essential for many fundamental cellular processes. Whether and how the internalization impacts cellular mechanics remains elusive. Whereas previous studies reported the contribution of the integrin activator, talin, in force development, the involvement of inhibitors is less documented. We identified ICAP-1 as an integrin inhibitor involved in mechanotransduction by co-working with NME2 to control clathrin-mediated endocytosis of integrins at the edge of focal adhesions (FA). Loss of ICAP-1 enables ß3-integrin-mediated force generation independently of ß1 integrin. ß3-integrin-mediated forces were associated with a decrease in ß3 integrin dynamics stemming from their reduced diffusion within adhesion sites and slow turnover of FA. The decrease in ß3 integrin dynamics correlated with a defect in integrin endocytosis. ICAP-1 acts as an adaptor for clathrin-dependent endocytosis of integrins. ICAP-1 controls integrin endocytosis by interacting with NME2, a key regulator of dynamin-dependent clathrin-coated pits fission. Control of clathrin-mediated integrin endocytosis by an inhibitor is an unprecedented mechanism to tune forces at FA.

Enterocytic differentiation is modulated by lipid rafts-dependent assembly of adherens junctions.

Integrity of the epithelial barrier is determined by apical junctional complexes which also participate in the signalling pathways inducing intestinal cell differentiation. Lipid rafts (LR) have been proposed to play a role in the organization and the function of these intercellular complexes. This study investigated potential mechanisms by which LR could participate in the establishment of adherens junctions (AJ) and the initiation of enterocytic differentiation. We showed that the differentiation of epithelial cells in rat colons correlates with the emergence of LR. Using HT-29 cells we demonstrated that during the differentiation process, LR are required for the recruitment and the association of p120ctn to E-cadherin. Silencing of flotillin-1, a LR component, alters the recruitment of AJ proteins in LR and delays the expression of differentiation markers. Furthermore, the ability of p120ctn/E-cadherin complexes to support cell differentiation is altered in HT-29 Rac1N17 cells. These results show a contributory role of LR in the enterocytic differentiation process, which serve as signalling platforms for Rac1-mediated organization of AJ. A better understanding of the mechanism involved in the establishment of junctional complex and their role in enterocytic differentiation provides new insights into the regulation of intestinal homeostasis.

CK2ß Is a Gatekeeper of Focal Adhesions Regulating Cell Spreading.

CK2 is a hetero-tetrameric serine/threonine protein kinase made up of two CK2α/α' catalytic subunits and two CK2ß regulatory subunits. The free CK2α subunit and the tetrameric holoenzyme have distinct substrate specificity profiles, suggesting that the spatiotemporal organization of the individual CK2 subunits observed in living cells is crucial in the control of the many cellular processes that are governed by this pleiotropic kinase. Indeed, previous studies reported that the unbalanced expression of CK2 subunits is sufficient to drive epithelial to mesenchymal transition (EMT), a process involved in cancer invasion and metastasis. Moreover, sub-stoichiometric expression of CK2ß compared to CK2α in a subset of breast cancer tumors was correlated with the induction of EMT markers and increased epithelial cell plasticity in breast carcinoma progression. Phenotypic changes of epithelial cells are often associated with the activation of phosphotyrosine signaling. Herein, using phosphotyrosine enrichment coupled with affinity capture and proteomic analysis, we show that decreased expression of CK2ß in MCF10A mammary epithelial cells triggers the phosphorylation of a number of proteins on tyrosine residues and promotes the striking activation of the FAK1-Src-PAX1 signaling pathway. Moreover, morphometric analyses also reveal that CK2ß loss increases the number and the spatial distribution of focal adhesion signaling complexes that coordinate the adhesive and migratory processes. Together, our findings allow positioning CK2ß as a gatekeeper for cell spreading by restraining focal adhesion formation and invasion of mammary epithelial cells.

Differential bioactivity of four BMP-family members as function of biomaterial stiffness.

While a soft film itself is not able to induce cell spreading, BMP-2 presented via such soft film (so called "matrix-bound BMP-2") was previously shown to trigger cell spreading, migration and downstream BMP-2 signaling. Here, we used thin films of controlled stiffness presenting matrix-bound BMPs to study the effect of four BMP members (BMP-2, 4, 7, 9) on cell adhesion and differentiation of skeletal progenitors. We performed automated high-content screening of cellular responses, including cell number, cell spreading area, SMAD phosphorylation and alkaline phosphatase activity. We revealed that the cell response to bBMPs is BMP-type specific, and involved certain BMP receptors and beta chain integrins. In addition, this response is stiffness-dependent for several receptors. The basolateral presentation of the BMPs allowed us to discriminate the specificity of cellular response, especiallyd the role of type I and II BMP receptors and of ß integrins in a BMP-type and stiffness-dependent manner. Notably, BMP-2 and BMP-4 were found to have distinct roles, while ALK5, previously known as a TGF-ß receptor was revealed to be involved in the BMP-pathway.

Integrin-based adhesion compartmentalizes ALK3 of the BMPRII to control cell adhesion and migration.

The spatial organization of cell-surface receptors is fundamental for the coordination of biological responses to physical and biochemical cues of the extracellular matrix. How serine/threonine kinase receptors, ALK3-BMPRII, cooperate with integrins upon BMP2 to drive cell migration is unknown. Whether the dynamics between integrins and BMP receptors intertwine in space and time to guide adhesive processes is yet to be elucidated. We found that BMP2 stimulation controls the spatial organization of BMPRs by segregating ALK3 from BMPRII into ß3 integrin-containing focal adhesions. The selective recruitment of ALK3 to focal adhesions requires ß3 integrin engagement and ALK3 activation. BMP2 controls the partitioning of immobilized ALK3 within and outside focal adhesions according to single-protein tracking and super-resolution imaging. The spatial control of ALK3 in focal adhesions by optogenetics indicates that ALK3 acts as an adhesive receptor by eliciting cell spreading required for cell migration. ALK3 segregation from BMPRII in integrin-based adhesions is a key aspect of the spatio-temporal control of BMPR signaling.

Interplay between integrins and cadherins to control bone differentiation upon BMP-2 stimulation.

Introduction: Upon BMP-2 stimulation, the osteoblastic lineage commitment in C2C12 myoblasts is associated with a microenvironmental change that occurs over several days. How does BMP-2 operate a switch in adhesive machinery to adapt to the new microenvironment and to drive bone cell fate is not well understood. Here, we addressed this question for BMP-2 delivered either in solution or physically bound of a biomimetic film, to mimic its presentation to cells via the extracellular matrix (ECM). Methods: Biommetics films were prepared using a recently developed automated method that enable high content studies of cellular processes. Comparative gene expressions were done using RNA sequencing from the encyclopedia of the regulatory elements (ENCODE). Gene expressions of transcription factors, beta chain (1, 3, 5) integrins and cadherins (M, N, and Cad11) were studied using quantitative PCR. ECM proteins and adhesion receptor expressions were also quantified by Western blots and dot blots. Their spatial organization in and around cells was studied using immuno-stainings. The individual effect of each receptor on osteogenic transcription factors and alkaline phosphatase expression were studied using silencing RNA of each integrin and cadherin receptor. The organization of fibronectin was studied using immuno-staining and quantitative microscopic analysis. Results: Our findings highlight a switch of integrin and cadherin expression during muscle to bone transdifferentiation upon BMP-2 stimulation. This switch occurs no matter the presentation mode, for BMP-2 presented in solution or via the biomimetic film. While C2C12 muscle cells express M-cadherin and Laminin-specific integrins, the BMP-2-induced transdifferentiation into bone cells is associated with an increase in the expression of cadherin-11 and collagen-specific integrins. Biomimetic films presenting matrix-bound BMP-2 enable the revelation of specific roles of the adhesive receptors depending on the transcription factor. Discussion: While ß3 integrin and cadherin-11 work in concert to control early pSMAD1,5,9 signaling, ß1 integrin and Cadherin-11 control RunX2, ALP activity and fibronectin organization around the cells. In contrast, while ß1 integrin is also important for osterix transcriptional activity, Cadherin-11 and ß5 integrin act as negative osterix regulators. In addition, ß5 integrin negatively regulates RunX2. Our results show that biomimetic films can be used to delinate the specific events associated with BMP-2-mediated muscle to bone transdifferentiation. Our study reveals how integrins and cadherins work together, while exerting distinct functions to drive osteogenic programming. Different sets of integrins and cadherins have complementary mechanical roles during the time window of this transdifferentiation.

Actin machinery and mechanosensitivity in invadopodia, podosomes and focal adhesions.

The invasiveness of cells is correlated with the presence of dynamic actin-rich membrane structures called invadopodia, which are membrane protrusions that are associated with localized polymerization of sub-membrane actin filaments. Similar to focal adhesions and podosomes, invadopodia are cell-matrix adhesion sites. Indeed, invadopodia share several features with podosomes, but whether they are distinct structures is still a matter of debate. Invadopodia are built upon an N-WASP-dependent branched actin network, and the Rho GTPase Cdc42 is involved in inducing invadopodial-membrane protrusion, which is mediated by actin filaments that are organized in bundles to form an actin core. Actin-core formation is thought to be an early step in invadopodium assembly, and the actin core is perpendicular to the extracellular matrix and the plasma membrane; this contrasts with the tangential orientation of actin stress fibers anchored to focal adhesions. In this Commentary, we attempt to summarize recent insights into the actin dynamics of invadopodia and podosomes, and the forces that are transmitted through these invasive structures. Although the mechanisms underlying force-dependent regulation of invadopodia and podosomes are largely unknown compared with those of focal adhesions, these structures do exhibit mechanosensitivity. Actin dynamics and associated forces might be key elements in discriminating between invadopodia, podosomes and focal adhesions. Targeting actin-regulatory molecules that specifically promote invadopodium formation is an attractive strategy against cancer-cell invasion.