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Adrenomedullin blockade induces regression of tumor neovessels through interference with vascular endothelial-cadherin signalling.

Khalfaoui-Bendriss G, Dussault N, Fernandez-Sauze S, Berenguer-Daizé C, Sigaud R, Delfino C, Cayol M, Metellus P, Chinot O, Mabrouk K, Martin PM, Ouafik L - Oncotarget (2015)

Bottom Line: At a molecular level, we show that AM blockade induces tyrosine phosphorylation of VE-cadherin at a critical tyrosine, Tyr731, which is sufficient to prevent the binding of β-catenin to the cytoplasmic tail of VE-cadherin leading to the inhibition of cell barrier function.Furthermore, we demonstrate activation of Src kinase by phosphorylation on Tyr416, supporting a role of Src to phosphorylate Tyr731-VE-cadherin.In this model, Src inhibition impairs αAM and αAMR-induced Tyr731-VE-cadherin phosphorylation in a dose-dependent manner, indicating that Tyr731-VE-cadherin phosphorylation state is dependent on Src activation.

View Article: PubMed Central - PubMed

Affiliation: Aix Marseille Université, CRO2, UMR_S 911, Faculté de Médecine, Marseille, France.

ABSTRACT
The cellular and molecular mechanisms by which adrenomedullin (AM) blockade suppresses tumor neovessels are not well defined. Herein, we show that AM blockade using anti-AM and anti-AM receptors antibodies targets vascular endothelial cells (ECs) and vascular smooth muscle cells (VSMCs), and induces regression of unstable nascent tumor neovessels. The underlying mechanism involved, and shown in vitro and in vivo in mice, is the disruption of the molecular engagement of the endothelial cell-specific junctional molecules vascular endothelial-cadherin (VE-cadherin)/β-catenin complex. AM blockade increases endothelial cell permeability by inhibiting cell-cell contacts predominantly through disruption of VE-cadherin/β-catenin/Akt signalling pathway, thereby leading to vascular collapse and regression of tumor neovessels. At a molecular level, we show that AM blockade induces tyrosine phosphorylation of VE-cadherin at a critical tyrosine, Tyr731, which is sufficient to prevent the binding of β-catenin to the cytoplasmic tail of VE-cadherin leading to the inhibition of cell barrier function. Furthermore, we demonstrate activation of Src kinase by phosphorylation on Tyr416, supporting a role of Src to phosphorylate Tyr731-VE-cadherin. In this model, Src inhibition impairs αAM and αAMR-induced Tyr731-VE-cadherin phosphorylation in a dose-dependent manner, indicating that Tyr731-VE-cadherin phosphorylation state is dependent on Src activation. We found that AM blockade induces β-catenin phosphorylation on Ser33/Ser37/Thr41 sites in both ECs and VSMCs both in vitro and in vivo in mice. These data suggest that AM blockade selectively induces regression of unstable tumor neovessels, through disruption of VE-cadherin signalling. Targeting AM system may present a novel therapeutic target to selectively disrupt assembly and induce regression of nascent tumor neovessels, without affecting normal stabilized vasculature.

No MeSH data available.


Related in: MedlinePlus

Morphogenetic activity of AM and αAMHUVECs (4 × 104 cells/well) were seeded into Matrigel-precoated wells and cultured in low-serum conditions (0.5% FCS) in presence of IgG control (70 μg/ml) (a), AM (10−8 M) (b), or αAM (70 μg/ml) (c). Photographs were taken 18 h later. Original magnification, × 20.
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Figure 2: Morphogenetic activity of AM and αAMHUVECs (4 × 104 cells/well) were seeded into Matrigel-precoated wells and cultured in low-serum conditions (0.5% FCS) in presence of IgG control (70 μg/ml) (a), AM (10−8 M) (b), or αAM (70 μg/ml) (c). Photographs were taken 18 h later. Original magnification, × 20.

Mentions: To explore the capacity of αAM and αAMR to inhibit angiogenesis, we measured their ability to disrupt the vascular structures formed by endothelial cells in an in vitro Matrigel assay. To this end, HUVECs that actively expressed AM and AM receptors and for which migration and invasion in Boyden assays in vitro were inhibited upon treatment with αAM or αAMR [29], were used as model to decipher the molecular mechanisms engaged by AM. Addition of AM (10−7 M) resulted in the formation of strong visible rings and cords of cells on growth factor-depleted Matrigel in the presence of 0.5% FCS (Figure 2b) when compared to non treated cells (Figure 2a). The incubation of HUVECs with αAM blocked tube formation and induced a clear cell-cell separation (Figure 2c) suggesting that AM promotes endothelial cell-cell interactions probably through modulation of junction proteins. Therefore, we tested the hypothesis that αAM and αAMR might interfere with VE-cadherin homophilic interactions.


Adrenomedullin blockade induces regression of tumor neovessels through interference with vascular endothelial-cadherin signalling.

Khalfaoui-Bendriss G, Dussault N, Fernandez-Sauze S, Berenguer-Daizé C, Sigaud R, Delfino C, Cayol M, Metellus P, Chinot O, Mabrouk K, Martin PM, Ouafik L - Oncotarget (2015)

Morphogenetic activity of AM and αAMHUVECs (4 × 104 cells/well) were seeded into Matrigel-precoated wells and cultured in low-serum conditions (0.5% FCS) in presence of IgG control (70 μg/ml) (a), AM (10−8 M) (b), or αAM (70 μg/ml) (c). Photographs were taken 18 h later. Original magnification, × 20.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4480698&req=5

Figure 2: Morphogenetic activity of AM and αAMHUVECs (4 × 104 cells/well) were seeded into Matrigel-precoated wells and cultured in low-serum conditions (0.5% FCS) in presence of IgG control (70 μg/ml) (a), AM (10−8 M) (b), or αAM (70 μg/ml) (c). Photographs were taken 18 h later. Original magnification, × 20.
Mentions: To explore the capacity of αAM and αAMR to inhibit angiogenesis, we measured their ability to disrupt the vascular structures formed by endothelial cells in an in vitro Matrigel assay. To this end, HUVECs that actively expressed AM and AM receptors and for which migration and invasion in Boyden assays in vitro were inhibited upon treatment with αAM or αAMR [29], were used as model to decipher the molecular mechanisms engaged by AM. Addition of AM (10−7 M) resulted in the formation of strong visible rings and cords of cells on growth factor-depleted Matrigel in the presence of 0.5% FCS (Figure 2b) when compared to non treated cells (Figure 2a). The incubation of HUVECs with αAM blocked tube formation and induced a clear cell-cell separation (Figure 2c) suggesting that AM promotes endothelial cell-cell interactions probably through modulation of junction proteins. Therefore, we tested the hypothesis that αAM and αAMR might interfere with VE-cadherin homophilic interactions.

Bottom Line: At a molecular level, we show that AM blockade induces tyrosine phosphorylation of VE-cadherin at a critical tyrosine, Tyr731, which is sufficient to prevent the binding of β-catenin to the cytoplasmic tail of VE-cadherin leading to the inhibition of cell barrier function.Furthermore, we demonstrate activation of Src kinase by phosphorylation on Tyr416, supporting a role of Src to phosphorylate Tyr731-VE-cadherin.In this model, Src inhibition impairs αAM and αAMR-induced Tyr731-VE-cadherin phosphorylation in a dose-dependent manner, indicating that Tyr731-VE-cadherin phosphorylation state is dependent on Src activation.

View Article: PubMed Central - PubMed

Affiliation: Aix Marseille Université, CRO2, UMR_S 911, Faculté de Médecine, Marseille, France.

ABSTRACT
The cellular and molecular mechanisms by which adrenomedullin (AM) blockade suppresses tumor neovessels are not well defined. Herein, we show that AM blockade using anti-AM and anti-AM receptors antibodies targets vascular endothelial cells (ECs) and vascular smooth muscle cells (VSMCs), and induces regression of unstable nascent tumor neovessels. The underlying mechanism involved, and shown in vitro and in vivo in mice, is the disruption of the molecular engagement of the endothelial cell-specific junctional molecules vascular endothelial-cadherin (VE-cadherin)/β-catenin complex. AM blockade increases endothelial cell permeability by inhibiting cell-cell contacts predominantly through disruption of VE-cadherin/β-catenin/Akt signalling pathway, thereby leading to vascular collapse and regression of tumor neovessels. At a molecular level, we show that AM blockade induces tyrosine phosphorylation of VE-cadherin at a critical tyrosine, Tyr731, which is sufficient to prevent the binding of β-catenin to the cytoplasmic tail of VE-cadherin leading to the inhibition of cell barrier function. Furthermore, we demonstrate activation of Src kinase by phosphorylation on Tyr416, supporting a role of Src to phosphorylate Tyr731-VE-cadherin. In this model, Src inhibition impairs αAM and αAMR-induced Tyr731-VE-cadherin phosphorylation in a dose-dependent manner, indicating that Tyr731-VE-cadherin phosphorylation state is dependent on Src activation. We found that AM blockade induces β-catenin phosphorylation on Ser33/Ser37/Thr41 sites in both ECs and VSMCs both in vitro and in vivo in mice. These data suggest that AM blockade selectively induces regression of unstable tumor neovessels, through disruption of VE-cadherin signalling. Targeting AM system may present a novel therapeutic target to selectively disrupt assembly and induce regression of nascent tumor neovessels, without affecting normal stabilized vasculature.

No MeSH data available.


Related in: MedlinePlus