RESUMO
Membrane overexpression of the receptor tyrosine kinase ErbB-2 (MErbB-2) accounts for a clinically aggressive breast cancer (BC) subtype (ErbB-2-positive) with increased incidence of metastases. We and others demonstrated that nuclear ErbB-2 (NErbB-2) also plays a key role in BC and is a poor prognostic factor in ErbB-2-positive tumors. The signal transducer and activator of transcription 3 (Stat3), another player in BC, has been recognized as a downstream mediator of MErbB-2 action in BC metastasis. Here, we revealed an unanticipated novel direction of the ErbB-2 and Stat3 interaction underlying BC metastasis. We found that Stat3 binds to its response elements (GAS) at the ErbB-2 promoter to upregulate ErbB-2 transcription in metastatic, ErbB-2-positive BC. We validated these results in several BC subtypes displaying metastatic and non-metastatic ability, highlighting Stat3 general role as upstream regulator of ErbB-2 expression in BC. Moreover, we showed that Stat3 co-opts NErbB-2 function by recruiting ErbB-2 as its coactivator at the GAS sites in the promoter of microRNA-21 (miR-21), a metastasis-promoting microRNA (miRNA). Using an ErbB-2 nuclear localization domain mutant and a constitutively activated ErbB-2 variant, we found that NErbB-2 role as a Stat3 coactivator and also its direct role as transcription factor upregulate miR-21 in BC. This reveals a novel function of NErbB-2 as a regulator of miRNAs expression. Increased levels of miR-21, in turn, downregulate the expression of the metastasis-suppressor protein programmed cell death 4 (PDCD4), a validated miR-21 target. Using an in vivo model of metastatic ErbB-2-postive BC, in which we silenced Stat3 and reconstituted ErbB-2 or miR-21 expression, we showed that both are downstream mediators of Stat3-driven metastasis. Supporting the clinical relevance of our results, we found an inverse correlation between ErbB-2/Stat3 nuclear co-expression and PDCD4 expression in ErbB-2-positive primary invasive BCs. Our findings identify Stat3 and NErbB-2 as novel therapeutic targets to inhibit ErbB-2-positive BC metastasis.
Assuntos
Proteínas Reguladoras de Apoptose/biossíntese , Neoplasias da Mama/genética , MicroRNAs/biossíntese , Proteínas de Ligação a RNA/biossíntese , Receptor ErbB-2/biossíntese , Fator de Transcrição STAT3/genética , Adolescente , Adulto , Idoso , Proteínas Reguladoras de Apoptose/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/genética , Pessoa de Meia-Idade , Metástase Neoplásica , Proteínas de Ligação a RNA/genética , Receptor ErbB-2/genética , Transdução de Sinais , Ativação Transcricional/genética , TransfecçãoRESUMO
LP07 is a new cell line derived from P07 lung tumor, spontaneously arisen in a BALB/c mouse. LP07 is composed of heterogeneous epithelioid polyhedric cells that proliferate at a slow rate, have low plating efficiency and are unable to grow in soft agar. Only some LP07 cells expressed cytokeratins while most of them were positive for vimentin. Ultrastructure studies showed that LP07 cells established rudimentary intercellular unions, formed glandular-like conducts and presented conspicuous secretory granules, suggesting an epithelial-glandular origin, with neuroendocrine components. Upon injection LP07 cells formed poorly differentiated non-invasive adenocarcinomas, and tumor bearing mice developed leukocytosis, hypercalcemia and cachexia. This tumor cell line constitutes a useful tool to study lung tumor biology and paraneoplastic syndromes.
Assuntos
Adenocarcinoma/patologia , Divisão Celular/fisiologia , Neoplasias Pulmonares/patologia , Síndromes Paraneoplásicas/patologia , Adenocarcinoma/sangue , Animais , Contagem de Células Sanguíneas , Peso Corporal , Cálcio/sangue , Testes de Carcinogenicidade , Adesão Celular , Linhagem da Célula , Movimento Celular , Cromossomos/genética , Análise Citogenética , Modelos Animais de Doenças , Feminino , Técnicas Imunoenzimáticas , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica , Transplante de Neoplasias , Síndromes Paraneoplásicas/sangue , Células Tumorais Cultivadas , Ativador de Plasminogênio Tipo Uroquinase/metabolismoRESUMO
The ability of tumor cells to respond to microenvironmental factors present in the target organ determines in part the successful development of a metastasis. In a previous work it was demonstrated that the conditioned medium (CM) from lungs of normal mice stimulates in vitro migration, proliferation and uPA activity of cells from a murine mammary adenocarcinoma moderately metastatic to lung. This CM also enhanced local and metastatic tumor growth. Here, we show that lung CM enhanced neovascularization when inoculated together with LM3 tumor cells into the skin of syngeneic mice. A similar tumor-induced angiogenesis response was obtained when lung CM was injected systemically. Western blot analysis of lung CM revealed the presence of some laminin fragments containing the sequence SIKVAV. To determine whether those molecules were responsible for the observed angiogenic effects, the CM was depleted of the peptides containing the SIKVAV sequence. We observed that the SIKVAV-depleted lung CM lost its ability to induce an enhancement of the tumor neovascular response. Our results suggest a role for the target organ in facilitating the neovascularization of tumor cells, probably through the participation of active peptides derived from the proteolytic degradation of the basement membrane component laminin.
