RESUMO
Tumor suppressive microRNAs are potent molecules that might cure cancer, one day. Despite the many advanced strategies for delivery of these microRNAs to the cell, there are few therapeutic microRNAs in clinical use. Progress in microRNA bioapplications is hindered by a high vulnerability of exogeneous microRNA molecules to RNase degradation that occurs in extra- and intracellular physiological conditions. In this proof-of-concept study, we use a programmable self-assembled DNA nanostructure bearing a "shuriken" shape to not only deliver but more importantly protect a tumor suppressive microRNA-145 for a sufficiently long time to exert its therapeutic effect in human colorectal cancer cells. Our DNA nanostructure harbored complementary sequences that can hybridize with the microRNA cargo. This brings the microRNA-DNA duplex very close to the core structure such that the microRNA cargo becomes sterically shielded from RNase's degradative activity. Our novel DNA nanostructure based protector concept removes the degradative bottleneck that may plague other nucleic acid delivery strategies and presents a new paradigm towards exploiting these microRNAs for anti-cancer therapy.
RESUMO
Epithelial-mesenchymal transition (EMT) is a crucial step in tumor progression, and the TGFß-SMAD signaling pathway as an inductor of EMT in many tumor types is well recognized. However, the role of non-canonical TGFß-TAK1 signaling in EMT remains unclear. Herein, we show that TAK1 deficiency drives metastatic skin squamous cell carcinoma earlier into EMT that is conditional on the elevated cellular ROS level. The expression of TAK1 is consistently reduced in invasive squamous cell carcinoma biopsies. Tumors derived from TAK1-deficient cells also exhibited pronounced invasive morphology. TAK1-deficient cancer cells adopt a more mesenchymal morphology characterized by higher number of focal adhesions, increase surface expression of integrin α5ß1 and active Rac1. Notably, these mutant cells exert an increased cell traction force, an early cellular response during TGFß1-induced EMT. The mRNA level of ZEB1 and SNAIL, transcription factors associated with mesenchymal phenotype is also upregulated in TAK1-deficient cancer cells compared with control cancer cells. We further show that TAK1 modulates Rac1 and RhoA GTPases activities via a redox-dependent downregulation of RhoA by Rac1, which involves the oxidative modification of low-molecular weight protein tyrosine phosphatase. Importantly, the treatment of TAK1-deficient cancer cells with Y27632, a selective inhibitor of Rho-associated protein kinase and antioxidant N-acetylcysteine augment and hinders EMT, respectively. Our findings suggest that a dysregulated balance in the activation of TGFß-TAK1 and TGFß-SMAD pathways is pivotal for TGFß1-induced EMT. Thus, TAK1 deficiency in metastatic cancer cells increases integrin:Rac-induced ROS, which negatively regulated Rho by LMW-PTP to accelerate EMT.
Assuntos
Transição Epitelial-Mesenquimal/efeitos dos fármacos , MAP Quinase Quinase Quinases/metabolismo , Neoplasias de Células Escamosas/enzimologia , Neoplasias de Células Escamosas/patologia , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Cutâneas/patologia , Animais , Fenômenos Biomecânicos/efeitos dos fármacos , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células HEK293 , Humanos , Integrina beta1/metabolismo , Integrina beta3/metabolismo , MAP Quinase Quinase Quinases/deficiência , Camundongos , Modelos Biológicos , Invasividade Neoplásica , Oxirredução/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Neoplasias Cutâneas/enzimologia , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Proteínas rac1 de Ligação ao GTP/metabolismo , Proteínas rho de Ligação ao GTP/metabolismoRESUMO
The use of nanomaterials has raised safety concerns, as their small size facilitates accumulation in and interaction with biological tissues. Here we show that exposure of endothelial cells to TiO2 nanomaterials causes endothelial cell leakiness. This effect is caused by the physical interaction between TiO2 nanomaterials and endothelial cells' adherens junction protein VE-cadherin. As a result, VE-cadherin is phosphorylated at intracellular residues (Y658 and Y731), and the interaction between VE-cadherin and p120 as well as ß-catenin is lost. The resulting signalling cascade promotes actin remodelling, as well as internalization and degradation of VE-cadherin. We show that injections of TiO2 nanomaterials cause leakiness of subcutaneous blood vessels in mice and, in a melanoma-lung metastasis mouse model, increase the number of pulmonary metastases. Our findings uncover a novel non-receptor-mediated mechanism by which nanomaterials trigger intracellular signalling cascades via specific interaction with VE-cadherin, resulting in nanomaterial-induced endothelial cell leakiness.