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Loss of phosphodiesterase 4D mediates acquired triapine resistance via Epac-Rap1-Integrin signaling.

ABSTRACT: Triapine, an anticancer thiosemicarbazone, is currently under clinical investigation. Whereas promising results were obtained in hematological diseases, trials in solid tumors widely failed. To understand mechanisms causing triapine insensitivity, we have analysed genomic alterations in a triapine-resistant SW480 subline (SW480/tria). Only one distinct genomic loss was observed specifically in SW480/tria cells affecting the phosphodiesterase 4D (PDE4D) gene locus. Accordingly, pharmacological inhibition of PDE4D resulted in significant triapine resistance in SW480 cells. Hence, we concluded that enhanced cyclic AMP levels might confer protection against triapine. Indeed, hyperactivation of both major downstream pathways, namely the protein kinase A (PKA)-cAMP response element-binding protein (Creb) and the exchange protein activated by cAMP (Epac)-Ras-related protein 1 (Rap1) signaling axes, was observed in SW480/tria cells. Unexpectedly, inhibition of PKA did not re-sensitize SW480/tria cells against triapine. In contrast, Epac activation resulted in distinct triapine resistance in SW480 cells. Conversely, knock-down of Epac expression and pharmacological inhibition of Rap1 re-sensitized SW480/tria cells against triapine. Rap1 is a well-known regulator of integrins. Accordingly, SW480/tria cells displayed enhanced plasma membrane expression of several integrin subunits, enhanced adhesion especially to RGD-containing matrix components, and bolstered activation/expression of the integrin downstream effectors Src and RhoA/Rac. Accordingly, integrin and Src inhibition resulted in potent triapine re-sensitization especially of SW480/tria cells. In summary, we describe for the first time integrin activation based on cAMP-Epac-Rap1 signaling as acquired drug resistance mechanism. combinations of triapine with inhibitors of several steps in this resistance cascade might be feasible strategies to overcome triapine insensitivity of solid tumors.

SUBMITTER: Miklos W

PROVIDER: S-EPMC5356681 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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Loss of phosphodiesterase 4D mediates acquired triapine resistance via Epac-Rap1-Integrin signaling.

Miklos Walter W Heffeter Petra P Pirker Christine C Hager Sonja S Kowol Christian R CR van Schoonhoven Sushilla S Stojanovic Mirjana M Keppler Bernhard K BK Berger Walter W

Oncotarget 20161201 51

Triapine, an anticancer thiosemicarbazone, is currently under clinical investigation. Whereas promising results were obtained in hematological diseases, trials in solid tumors widely failed. To understand mechanisms causing triapine insensitivity, we have analysed genomic alterations in a triapine-resistant SW480 subline (SW480/tria). Only one distinct genomic loss was observed specifically in SW480/tria cells affecting the phosphodiesterase 4D (PDE4D) gene locus. Accordingly, pharmacological in ...[more]

PMID: 27602951

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Project description:Decellularized tissues offer a unique tool for developing regenerative biomaterials or in vitro platforms for the study of cell-extracellular matrix (ECM) interactions. One main challenge associated with decellularized lung tissue is that ECM components can be stripped away or altered by the detergents used to remove cellular debris. Without characterizing the composition of lung decellularized ECM (dECM) and the cellular response caused by the altered composition, it is difficult to utilize dECM for regeneration and specifically, engineering the complexities of the alveolar-capillary barrier. This study takes steps towards uncovering if dECM must be enhanced with lost ECM proteins to achieve proper epithelial barrier formation. To achieve this, the epithelial barrier function was assessed on dECM coatings with and without the systematic addition of several key basement membrane proteins. After comparing barrier function on collagen I, fibronectin, laminin, and dECM in varying combinations as an in vitro coating, the alveolar epithelium exhibited superior barrier function when dECM was supplemented with laminin as evidenced by trans-epithelial electrical resistance (TEER) and permeability assays. Increased barrier resistance with laminin addition was associated with upregulation of Claudin-18, E-cadherin, and junction adhesion molecule (JAM)-A, and stabilization of zonula occludens (ZO)-1 at junction complexes. The Epac/Rap1 pathway was observed to play a role in the ECM-mediated barrier function determined by protein expression and Epac inhibition. These findings revealed potential ECM coatings and molecular therapeutic targets for improved regeneration with decellularized scaffolds. STATEMENT OF SIGNIFICANCE: Efforts to produce a transplantable organ-scale biomaterial for lung regeneration has not been entirely successful to date, due to incomplete cell-cell junction formation, ultimately leading to severe edema in vivo. To fully understand the process of alveolar junction formation on ECM-derived biomaterials, this research has characterized and tailored decellularized ECM (dECM) to mitigate reductions in barrier strength or cell attachment caused by abnormal ECM compositions or detergent damage to dECM. These results indicate that laminin-driven Epac signaling plays a vital role in the stabilization of the alveolar barrier. Addition of laminin or Epac agonists during alveolar regeneration can reduce epithelial permeability within bioengineered lungs.

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Dataset Information

Loss of phosphodiesterase 4D mediates acquired triapine resistance via Epac-Rap1-Integrin signaling.

Publications

Loss of phosphodiesterase 4D mediates acquired triapine resistance via Epac-Rap1-Integrin signaling.

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