RESUMEN
Tumor cell migration has a fundamental role in early steps of metastasis, the fatal hallmark of cancer. In the present study, we investigated the effects of the tyrosine phosphatase, SRC-homology 2 domain-containing phosphatase 2 (SHP2), on cell migration in metastatic triple-negative breast cancer (TNBC), an aggressive disease associated with a poor prognosis for which a targeted therapy is not yet available. Using mouse models and multiphoton intravital imaging, we have identified a crucial effect of SHP2 on TNBC cell motility in vivo. Further, analysis of TNBC cells revealed that SHP2 also influences cell migration, chemotaxis and invasion in vitro. Unbiased phosphoproteomics and biochemical analysis showed that SHP2 activates several SRC-family kinases and downstream targets, most of which are inducers of migration and invasion. In particular, direct interaction between SHP2 and c-SRC was revealed by a fluorescence resonance energy transfer assay. These results suggest that SHP2 is a crucial factor during early steps of TNBC migration to distant organs.
Asunto(s)
Neoplasias de la Mama/enzimología , Proteínas de Neoplasias/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 11/metabolismo , Familia-src Quinasas/metabolismo , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Movimiento Celular , Femenino , Xenoinjertos , Humanos , Ratones , Invasividad Neoplásica , Proteínas de Neoplasias/genética , Trasplante de Neoplasias , Proteína Tirosina Fosfatasa no Receptora Tipo 11/genética , Familia-src Quinasas/genéticaRESUMEN
Protein tyrosine phosphatase alpha (PTPα/PTPRA) was shown previously to be overexpressed in human primary breast cancers, and to suppress apoptosis in estrogen receptor-negative breast cancer cells in vitro. However, it is not known whether PTPα is important for mammary tumor initiation, maintenance and/or progression. We have used a combination of three-dimensional cultures, a transgenic mouse model of breast cancer lacking PTPα as well as xenografts of human breast cancer cell lines to address these questions. We found that PTPα knockdown after overt tumor development reduced the growth of HER2-positive human breast cancer cell lines, and that this effect was accompanied by a reduction in AKT phosphorylation. However, PTPα knockdown did not affect invasiveness of HER2-positive human breast cancer cells grown in three-dimensional cultures. Moreover, in MMTV-NeuNT/PTPα(-/-) mice, PTPα ablation did not affect NeuNT-evoked tumor onset or metastasis but decreased the number of tumors per mouse. Thus, we demonstrate that PTPα contributes to both HER2/Neu-mediated mammary tumor initiation and maintenance. Our results suggest that inhibition of PTPα can have a beneficial effect on HER2-positive breast cancers, but that inhibition of additional targets is needed to block breast tumorigenesis.
Asunto(s)
Neoplasias de la Mama/genética , Interferencia de ARN , Receptor ErbB-2/genética , Proteínas Tirosina Fosfatasas Clase 4 Similares a Receptores/genética , Animales , Neoplasias de la Mama/patología , Neoplasias de la Mama/terapia , Técnicas de Cultivo de Célula , Línea Celular , Línea Celular Tumoral , Humanos , Immunoblotting , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundario , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/metabolismo , Neoplasias Mamarias Experimentales/patología , Ratones , Ratones Noqueados , Ratones Transgénicos , ARN Interferente Pequeño/genética , Receptor ErbB-2/metabolismo , Proteínas Tirosina Fosfatasas Clase 4 Similares a Receptores/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Malignant mesothelioma is an asbestos-related, aggressive tumour, resistant to most anticancer therapies. Akt is a key mediator of mesothelioma cell survival and chemoresistance. This study aimed to clarify the mechanism by which taurolidine (TN), a known synthetic compound with antimicrobial and antineoplastic properties, leads to mesothelioma cell death. Apoptosis was studied by annexin V binding, cell cycle analysis, caspase-8 activation, poly(ADP-ribose) polymerase (PARP) cleavage and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labelling (TUNEL). Oxidative stress was measured by nitrite production and DNA oxidative damage. Protein expression and phosphorylation were evaluated by immunoprecipitation and immunoblotting. TN induces cell death of mesothelioma cells, but not of non-neoplastic human mesothelial cells. After TN treatment of mesothelioma cells, Akt but not extracellular signal-regulated kinase (Erk) 1/2 activity is inhibited a in time- and dose-dependent manner. Protein phosphatase (PP)1alpha and PP2A are activated several hours after drug addition. Apoptosis induced by TN is driven by oxidative stress and cell exposure to sulfydryl donors, such as glutathione monoethylester and l-N-acetylcysteine, significantly reduced pro-apoptotic effects and Akt inhibition. Conversely, expression of constitutively activated Akt did not affect cytoxicity elicited by TN, which retained its ability to inhibit the kinase. TN induces mesothelioma cell death via oxidative stress, accompanied by inhibition of Akt signalling. This provides a promising molecular rationale for TN as local treatment of malignant mesothelioma.