Regulation of urokinase receptor function and pericellular proteolysis by the integrin α5β1

Bass, Rosemary; Ellis, Vincent

doi:10.1160/th08-08-0558

Cited by 14 publications

(8 citation statements)

References 44 publications

(62 reference statements)

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“…Our previous studies using both unbiased and biased approaches demonstrated that ATN-658 inhibits integrin signaling in vitro and in vivo as well as co-localization of α5β1 and uPAR [22], [27]. Our data suggest that the effects of ATN-658 on perturbing the uPAR-α5β1 interaction is indirect and direct binding of uPAR to integrin α5β1 or any other integrin using purified components has not been demonstrated [57], suggesting that these interactions require multiple components present at the cell surface. The existence of a signalosome containing uPAR has been postulated by D'Allessio and Blasi [58] and the data obtained to date supports the hypothesis that ATN-658 disrupts the signalosome leading to global effects on integrin signaling and inhibition of tumor progression (Fig.…”

Section: Discussionmentioning

confidence: 59%

Identification of a New Epitope in uPAR as a Target for the Cancer Therapeutic Monoclonal Antibody ATN-658, a Structural Homolog of the uPAR Binding Integrin CD11b (αM)

Yuan

Wei

et al. 2014

PLoS ONE

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The urokinase plasminogen activator receptor (uPAR) plays a role in tumor progression and has been proposed as a target for the treatment of cancer. We recently described the development of a novel humanized monoclonal antibody that targets uPAR and has anti-tumor activity in multiple xenograft animal tumor models. This antibody, ATN-658, does not inhibit ligand binding (i.e. uPA and vitronectin) to uPAR and its mechanism of action remains unclear. As a first step in understanding the anti-tumor activity of ATN-658, we set out to identify the epitope on uPAR to which ATN-658 binds. Guided by comparisons between primate and human uPAR, epitope mapping studies were performed using several orthogonal techniques. Systematic site directed and alanine scanning mutagenesis identified the region of aa 268–275 of uPAR as the epitope for ATN-658. No known function has previously been attributed to this epitope Structural insights into epitope recognition were obtained from structural studies of the Fab fragment of ATN-658 bound to uPAR. The structure shows that the ATN-658 binds to the DIII domain of uPAR, close to the C-terminus of the receptor, corroborating the epitope mapping results. Intriguingly, when bound to uPAR, the complementarity determining region (CDR) regions of ATN-658 closely mimic the binding regions of the integrin CD11b (αM), a previously identified uPAR ligand thought to be involved in leukocyte rolling, migration and complement fixation with no known role in tumor progression of solid tumors. These studies reveal a new functional epitope on uPAR involved in tumor progression and demonstrate a previously unrecognized strategy for the therapeutic targeting of uPAR.

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Section: Discussionmentioning

confidence: 59%

Identification of a New Epitope in uPAR as a Target for the Cancer Therapeutic Monoclonal Antibody ATN-658, a Structural Homolog of the uPAR Binding Integrin CD11b (αM)

Yuan

Wei

et al. 2014

PLoS ONE

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“…When necessary these cells were differentiated by culturing in Medium 231 supplemented with 1% fetal bovine serum and heparin (30 g/ml) for 7 days. ␣5CHO (clone 17) and B2CHO cell lines were as previously detailed (29) and HT1080 were obtained from ATCC. These cells were maintained in Dulbecco's modified Eagle's medium (DMEM), supplemented with 10% (v/v) fetal calf serum, and 0.1 mM nonessential amino acids.…”

Section: Methodsmentioning

confidence: 99%

Binding of Extracellular Maspin to β1 Integrins Inhibits Vascular Smooth Muscle Cell Migration

Bass

Wagstaff

Ravenhill

et al. 2009

Journal of Biological Chemistry

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Maspin is a serpin that has multiple effects on cell behavior, including inhibition of migration. How maspin mediates these diverse effects remains unclear, as it is devoid of protease inhibitory activity. We have previously shown that maspin rapidly inhibits the migration of vascular smooth muscle cells (VSMC), suggesting the involvement of direct interactions with cell surface proteins. Here, using immunofluorescence microscopy, we demonstrate that maspin binds specifically to the surface of VSMC in the dedifferentiated, but not the differentiated, phenotype. Ligand blotting of VSMC lysates revealed the presence of several maspin-binding proteins, with a protein of 150 kDa differentially expressed between the two VSMC phenotypes. Western blotting suggested that this protein was the ␤1 integrin subunit, and subsequently both ␣3␤1 and ␣5␤1, but not ␣v␤3, were shown to associate with maspin by coimmunoprecipitation. Specific binding of these integrins was also observed using maspin-affinity chromatography, using HT1080 cell lysates. Direct binding of maspin to ␣5␤1 was confirmed using a recombinant ␣5␤1-Fc fusion protein. Using conformation-dependent anti-␤1 antibodies, maspin binding to VSMC was found to lead to a decrease in the activation status of the integrin. The functional involvement of ␣5␤1 in mediating the effect of maspin was established by the inhibition of migration of CHO cells overexpressing human ␣5 integrin, but not those lacking ␣5 expression. Our observations suggest that maspin engages in specific interactions with a limited number of integrins on VSMC, leading to their inactivation, and that these interactions are responsible for the effects of maspin in the pericellular environment.Maspin is a member of the serpin family of serine protease inhibitors (SERPINB5).2 It was originally identified as a gene down-regulated in invasive breast cancer and proposed as a class II tumor suppressor (1), and has since been shown to have many effects on cellular behavior that are consistent with this activity. It has been shown to decrease the proliferation, migration, and metastasis of tumor cells in vivo (1, 2) and their invasion in vitro (3, 4), and to increase apoptosis of endothelial cells (5) and inhibit angiogenesis (6). However, the cellular effects of maspin are not restricted to tumor cells, and we have demonstrated that maspin can inhibit the migration of vascular smooth muscle cells (7).VSMC migration is a key event in the development of atherosclerosis (8), and contributes significantly to restenosis after angioplasty (9) and transplant arteriosclerosis (10). VSMC are not terminally differentiated and acquire migratory capacity as part of a phenotypic switch from a contractile, quiescent state to a dedifferentiated phenotype, characterized by proliferation and increased extracellular matrix synthesis, in addition to motility (11). This allows VSMC to respond to environmental cues following vascular injury. The phenotypic plasticity of VSMC is regulated by an array of signals, among which integr...

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“…Integrins link ECM to the actin cytoskeleton through FAK/ILK/SFK/Rho proteins pathway providing the traction necessary for cell motility (Geiger et al, 2001). Integrins regulate the localization and the activity of urokinase-type plasminogen activator (uPA)/uPA receptor (Ghosh et al, 2000; Wei et al, 2007; Bass and Ellis, 2009) and matrix metalloproteinases (MMPs; Lamar et al, 2008; Morozevich et al, 2009) therefore controlling ECM remodeling and the invasive process.…”

Section: Integrins In Gliomamentioning

confidence: 99%

Integrins and p53 pathways in glioblastoma resistance to temozolomide

Martin¹,

Janoušková²,

Dontenwill³

2012

Front. Oncol.

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Glioblastoma is the most common malignant primary brain tumor. Surgical resection, postoperative radiotherapy plus concomitant and adjuvant chemotherapy with temozolomide (TMZ) is the standard of care for newly diagnosed glioblastoma. In the past decade, efforts have been made to decipher genomic and core pathway alterations to identify clinically relevant glioblastoma subtypes. Based on these studies and more academic explorations, new potential therapeutic targets were found and several targeting agents were developed. Such molecules should hopefully overcome the resistance of glioblastoma to the current therapy. One of the hallmarks of glioblastoma subtypes was the enrichment of extracellular matrix/invasion-related genes. Integrins, which are cell adhesion molecules important in glioma cell migration/invasion and angiogenesis were one of those genes. Integrins seem to be pertinent therapeutic targets and antagonists recently reached the clinic. Although the p53 pathway appears often altered in glioblastoma, conflicting results can be found in the literature about the clinically relevant impact of the p53 status in the resistance to TMZ. Here, we will summarize the current knowledge on (1) integrin expression, (2) p53 status, and (3) relationship between integrins and p53 to discuss their potential impact on the resistance of glioblastoma to temozolomide.

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Regulation of urokinase receptor function and pericellular proteolysis by the integrin α5β1

Cited by 14 publications

References 44 publications

Identification of a New Epitope in uPAR as a Target for the Cancer Therapeutic Monoclonal Antibody ATN-658, a Structural Homolog of the uPAR Binding Integrin CD11b (αM)

Identification of a New Epitope in uPAR as a Target for the Cancer Therapeutic Monoclonal Antibody ATN-658, a Structural Homolog of the uPAR Binding Integrin CD11b (αM)

Binding of Extracellular Maspin to β1 Integrins Inhibits Vascular Smooth Muscle Cell Migration

Integrins and p53 pathways in glioblastoma resistance to temozolomide

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