RESUMO
Brain metastasis in breast cancer remains difficult to treat and its incidence is increasing. Therefore, the development of new therapies is of utmost clinical relevance. Recently, toll-like receptor (TLR) 4 was correlated with IL6 expression and poor prognosis in 1 215 breast cancer primaries. In contrast, we demonstrated that TLR4 stimulation reduces microglia-assisted breast cancer cell invasion. However, the expression, prognostic value, or therapeutic potential of TLR signaling in breast cancer brain metastasis have not been investigated. We thus tested the prognostic value of various TLRs in two brain-metastasis gene sets. Furthermore, we investigated different TLR agonists, as well as MyD88 and TRIF-deficient microenvironments in organotypic brain-slice ex vivo co-cultures and in vivo colonization experiments. These experiments underline the ambiguous roles of TLR4, its adapter MyD88, and the target nitric oxide (NO) during brain colonization. Moreover, analysis of the gene expression datasets of breast cancer brain metastasis patients revealed associations of TLR1 and IL6 with poor overall survival. Finally, our finding that a single LPS application at the onset of colonization shapes the later microglia/macrophage reaction at the macro-metastasis brain-parenchyma interface (MMPI) and reduces metastatic infiltration into the brain parenchyma may prove useful in immunotherapeutic considerations.
Assuntos
Neoplasias Encefálicas , Neoplasias da Mama , Humanos , Feminino , Receptor 4 Toll-Like/metabolismo , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Interleucina-6/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias da Mama/genética , Encéfalo/patologia , Neoplasias Encefálicas/tratamento farmacológico , Proteínas Adaptadoras de Transporte Vesicular/metabolismo , Microambiente TumoralRESUMO
More than half of all brain metastases show infiltrating rather than displacing growth at the macro-metastasis/organ parenchyma interface (MMPI), a finding associated with shorter survival. The lymphoid enhancer-binding factor-1 (LEF1) is an epithelial-mesenchymal transition (EMT) transcription factor that is commonly overexpressed in brain-colonizing cancer cells. Here, we overexpressed LEF1 in an in vivo breast cancer brain colonization model. It shortened survival, albeit without engaging EMT at the MMPI. By differential proteome analysis, we identified a novel function of LEF1 as a regulator of the glutathione (GSH) system, the principal cellular redox buffer. LEF1 overexpression also conferred resistance against therapeutic GSH depletion during brain colonization and improved management of intracellular ROS. We conclude that besides EMT, LEF1 facilitates metastasis by improving the antioxidative capacity of epithelial breast cancer cells, in particular during colonization of the brain parenchyma.
Assuntos
Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/secundário , Neoplasias da Mama/patologia , Glutationa/metabolismo , Fator 1 de Ligação ao Facilitador Linfoide/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Encéfalo/patologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Transição Epitelial-Mesenquimal/fisiologia , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Tecido Parenquimatoso/patologiaRESUMO
Mutations and activation of the PI3K signaling pathway in breast cancer cells have been linked to brain metastases. However, here we describe that in some breast cancer brain metastases samples the protein expression of PI3K signaling components is restricted to the metastatic microenvironment. In contrast to the therapeutic effects of PI3K inhibition on the breast cancer cells, the reaction of the brain microenvironment is less understood. Therefore we aimed to quantify the PI3K pathway activity in breast cancer brain metastasis and investigate the effects of PI3K inhibition on the central nervous system (CNS) microenvironment. First, to systematically quantify the PI3K pathway activity in breast cancer brain metastases, we performed a prospective biomarker study using a reverse phase protein array (RPPA). The majority, namely 30 out of 48 (62.5%) brain metastatic tissues examined, revealed high PI3K signaling activity that was associated with a median overall survival (OS) of 9.41 months, while that of patients, whose brain metastases showed only moderate or low PI3K activity, amounted to only 1.93 and 6.71 months, respectively. Second, we identified PI3K as a master regulator of metastasis-promoting macrophages/microglia during CNS colonization; and treatment with buparlisib (BKM120), a pan-PI3K Class I inhibitor with a good blood-brain-barrier penetrance, reduced their metastasis-promoting features. In conclusion, PI3K signaling is active in the majority of breast cancer brain metastases. Since PI3K inhibition does not only affect the metastatic cells but also re-educates the metastasis-promoting macrophages/microglia, PI3K inhibition may hold considerable promise in the treatment of brain metastasis and the respective microenvironment.
Assuntos
Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/secundário , Neoplasias da Mama/patologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Macrófagos/enzimologia , Microglia/enzimologia , Adulto , Idoso , Aminopiridinas/uso terapêutico , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Proteínas dos Microfilamentos/metabolismo , Microglia/efeitos dos fármacos , Pessoa de Meia-Idade , Morfolinas/uso terapêutico , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismoRESUMO
PURPOSE: Calcium ions are highly versatile spacial and temporal intracellular signals of non-excitable cells and have an important impact on nearly every aspect of cellular life controlling cell growth, metabolism, fluid secretion, information processing, transcription, apoptosis, and motility. Neurons and glia respond to stimuli, including neurotransmitters, neuromodulators, and hormones, which increase the intracellular calcium concentration. The function of intracellular calcium in gliomas is unknown. Lots of daily used drugs may act via receptors that can be linked to the intracellular calcium system and therefore could influence glioma biology. METHODS: Glioma cells were loaded with the calcium ion sensitive dye Fura 2-AM. Subsequently, cells were stimulated with 25 different medical drugs for 30 s. The increase of free intracellular calcium ions was measured and calculated by a microscope-camera-computer-unit. RESULTS: Except for the buffer solution HEPES that served as negative control and for the cortisol derivative dexamethasone, all other 24 tested drugs induced a rise of intracellular calcium ions. The cellular calcium responses were classified into seven functional groups. The tested substances activated several types of calcium channels and receptors. CONCLUSIONS: Our study impressively demonstrates that medical drugs are potent inducers of intracellular calcium signals. Totally unexpected, the results show a high amount of functional cellular receptors and channels on glioma cells, which could be responsible for certain biological effects like migration and cell growth. This calcium imaging study proves the usability of the calcium imaging as a screening system for functional receptors on human glioma cells.
Assuntos
Membrana Celular/metabolismo , Glioblastoma/metabolismo , Medicamentos sem Prescrição/farmacologia , Receptores de Superfície Celular/agonistas , Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Diagnóstico por Imagem/instrumentação , Diagnóstico por Imagem/métodos , Humanos , Modelos Biológicos , Medicamentos sem Prescrição/classificação , Receptores de Superfície Celular/metabolismo , Receptores de Superfície Celular/fisiologia , Células Tumorais CultivadasRESUMO
The course of autoimmune inflammatory diseases of the central nervous system (CNS) can be influenced by infections. Here we assessed the disease-modulating effects of the most frequent respiratory pathogen Streptococcus pneumonia on the course of experimental autoimmune encephalomyelitis (EAE). Mice were immunized with myelin oligodendrocyte glycoprotein 35-55 (MOG(35-55)) peptide, challenged intraperitoneally with live S. pneumoniae type 3, and then treated with ceftriaxone. EAE was monitored by a clinical score for 35 days after immunization. EAE was unaltered in mice infected with S. pneumoniae 2 days before and 21 days after the first MOG(35-55) injection but was more severe in animals infected 7 days after the first MOG(35-55) injection. The antigen-driven systemic T-cell response was unaltered, and the intraspinal Th1 cytokine mRNA concentrations at the peak of disease were unchanged. The composition of CNS-infiltrating cells and subsequent tissue destruction were only slightly increased after S. pneumoniae infection. In contrast, the serum levels of tumor necrosis factor alpha and interleukin-6 and spinal interleukin-6 levels were elevated, and the expression of major histocompatibility complex class II molecules, CD80, and CD86 on splenic dendritic cells were enhanced early after infection. Serum cytokine concentrations were not elevated, and EAE was not aggravated by S. pneumoniae infection in Toll-like receptor 2 (TLR2)-deficient mice. In conclusion, infection with S. pneumoniae worsens EAE probably by elevation of proinflammatory cytokines and activation of dendritic cells in the systemic circulation via TLR2 and cross talk through the blood-brain barrier.