RESUMO
Oncocytomas represent a subset of benign pituitary adenomas that are characterized by significant mitochondrial hyperplasia. Mitochondria are key organelles for energy generation and metabolic intermediate production for biosynthesis in tumour cells, so understanding the mechanism underlying mitochondrial biogenesis and its impact on cellular metabolism in oncocytoma is vital. Here, we studied surgically resected pituitary oncocytomas by using multi-omic analyses. Whole-exome sequencing did not reveal any nuclear mutations, but identified several somatic mutations of mitochondrial DNA, and dysfunctional respiratory complex I. Metabolomic analysis suggested that oxidative phosphorylation was reduced within individual mitochondria, and that there was no reciprocal increase in glycolytic activity. Interestingly, we found a reduction in the cellular lactate level and reduced expression of lactate dehydrogenase A (LDHA), which contributed to mitochondrial biogenesis in an in vitro cell model. It is of note that the hypoxia-response signalling pathway was not upregulated in pituitary oncocytomas, thereby failing to enhance glycolysis. Proteomic analysis showed that 14-3-3η was exclusively overexpressed in oncocytomas, and that 14-3-3η was capable of inhibiting glycolysis, leading to mitochondrial biogenesis in the presence of rotenone. In particular, 14-3-3η inhibited LDHA by direct interaction in the setting of complex I dysfunction, highlighting the role of 14-3-3η overexpression and inefficient oxidative phosphorylation in oncocytoma mitochondrial biogenesis. These findings deepen our understanding of the metabolic changes that occur within oncocytomas, and shine a light on the mechanism of mitochondrial biogenesis, providing a novel perspective on metabolic adaptation in tumour cells. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.
Assuntos
Proteínas 14-3-3/metabolismo , Adenoma Oxífilo/enzimologia , Metabolismo Energético , L-Lactato Desidrogenase/metabolismo , Mitocôndrias/enzimologia , Biogênese de Organelas , Neoplasias Hipofisárias/enzimologia , Proteínas 14-3-3/genética , Adenoma Oxífilo/genética , Adenoma Oxífilo/patologia , Adulto , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Complexo I de Transporte de Elétrons/metabolismo , Feminino , Glicólise , Células HEK293 , Células HeLa , Humanos , L-Lactato Desidrogenase/genética , Masculino , Pessoa de Meia-Idade , Mitocôndrias/patologia , Mutação , Fosforilação Oxidativa , Neoplasias Hipofisárias/genética , Neoplasias Hipofisárias/patologia , Transdução de Sinais , Microambiente TumoralRESUMO
A common hallmark of amyloids is their resistance to an array of proteases, highlighting the difficulty in degrading these disease-related aggregated proteinaceous materials. Here, we report on the potent activity of cathepsin L (CtsL), a lysosomal protease that proteolyzes the Parkinson's disease-related amyloid formed by α-synuclein (α-syn). Using liquid chromatography with mass spectrometry and transmission electron microscopy, an elegant mechanism is revealed on the residue and ultrastructural level, respectively. Specifically, CtsL always truncates α-syn fibrils first at the C-terminus before attacking the internal ß-sheet-rich region between residues 30 and 100. This suggests that only upon removal of the α-syn C-terminus can CtsL gain access to residues within the amyloid core. Interestingly, three of the four mapped sites contain a glycine residue (G36, G41, and G51) that is likely to be involved in a ß-turn in the fibril, whereupon cutting would lead to solvent exposure of internal residues and allow further proteolysis. Via close inspection of the fibril morphology, products resulting from CtsL degradation show imperfections along the fibril axis, with missing protein density as though they have been cannibalized. The ability of CtsL to degrade α-syn amyloid fibrils offers a promising strategy for improving the cellular clearance of aggregated α-syn through the modulation of protease levels and activity.
Assuntos
Amiloide/metabolismo , Catepsina L/metabolismo , alfa-Sinucleína/metabolismo , Acetilação , Amiloide/ultraestrutura , Catepsina L/genética , Cromatografia Líquida de Alta Pressão , Glicina/química , Humanos , Concentração de Íons de Hidrogênio , Cinética , Microscopia Eletrônica de Transmissão , Peso Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Mapeamento de Peptídeos , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Processamento de Proteína Pós-Traducional , Estabilidade Proteica , Proteólise , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , alfa-Sinucleína/química , alfa-Sinucleína/genéticaRESUMO
Amyloid is a systemic disease characterized by extracellular deposition of misfolded protein. Gastrointestinal and peritoneal deposition of light chain (AL) amyloid is an under-recognized manifestation of this systemic disease, usually as a late sequela. Here we present a case of recently diagnosed AL peritoneal amyloid that presented in the context of recurrent, acute onset abdominal discomfort and was found to have bowel obstruction complicated by perforation in the setting of AL-mediated gastrointestinal tract infiltration and dysmotility.
RESUMO
We present a case of a patient who presented to the emergency department with symptoms suggestive of a chronic obstructive pulmonary disease (COPD) exacerbation one day after receiving the BNT162b2 COVID-19 booster vaccine. Laboratory studies indicated she was in a state of inflammation, but not infection. Other potential triggers, including cardiac and viral etiologies, were ruled out. To our knowledge, this is the first documented case of a COPD exacerbation following the administration of the BNT162b2 COVID-19 vaccine.
RESUMO
Immunotherapy is a promising new therapeutic field that has demonstrated significant benefits in many solid-tumor malignancies, such as metastatic melanoma and non-small cell lung cancer. However, only a subset of these patients responds to treatment. Glioblastoma (GBM) is the most common malignant primary brain tumor with a poor prognosis of 14.6 months and few treatment advancements over the last 10 years. There are many clinical trials testing immune therapies in GBM, but patient responses in these studies have been highly variable and a definitive benefit has yet to be identified. Biomarkers are used to quantify normal physiology and physiological response to therapies. When extensively characterized and vigorously validated, they have the potential to delineate responders from non-responders for patients treated with immunotherapy in malignancies outside of the central nervous system (CNS) as well as GBM. Due to the challenges of current modalities of radiographic diagnosis and disease monitoring, identification of new predictive and prognostic biomarkers to gauge response to immune therapy for patients with GBM will be critical in the precise treatment of this highly heterogenous disease. This review will explore the current and future strategies for the identification of potential biomarkers in the field of immunotherapy for GBM, as well as highlight major challenges of adapting immune therapy for CNS malignancies.
Assuntos
Biomarcadores/metabolismo , Neoplasias Encefálicas/imunologia , Glioblastoma/imunologia , Imunoterapia/métodos , Neoplasias Encefálicas/patologia , Glioblastoma/patologia , HumanosRESUMO
Preclinical models that reliably recapitulate the immunosuppressive properties of human gliomas are essential to assess immune-based therapies. GL261 murine glioma cells are widely used as a syngeneic animal model of glioma, however, it has become common practice to transfect these cells with luciferase for fluorescent tumor tracking. The aim of this study was to compare the survival of mice injected with fluorescent or non-fluorescent GL261 cells and characterize the differences in their tumor microenvironment. Mice were intracranially implanted with GL261, GL261 Red-FLuc or GL261-Luc2 cells at varying doses. Cytokine profiles were evaluated by proteome microarray and Kaplan-Meier survival analysis was used to determine survival differences. Median survival for mice implanted with 5 × 104 GL261 cells was 18 to 21 days. The GL261 Red-FLuc implanted mice cells did not reach median survival at any tumor dose. Mice injected with 3 × 105 GL261-Luc2 cells reached median survival at 23 days. However, median survival was significantly prolonged to 37 days in mice implanted with 5 × 104 GL261-Luc2 cells. Additionally, proteomic analyses revealed significantly elevated inflammatory cytokines in the supernatants of the GL261 Red-FLuc cells and GL261-Luc2 cells. Our data suggest that GL261 Red-FLuc and GL261-Luc2 murine models elicit an anti-tumor immune response by increasing pro-inflammatory modulators.
Assuntos
Neoplasias Encefálicas/metabolismo , Citocinas/metabolismo , Glioma/metabolismo , Luciferases/imunologia , Regulação para Cima , Animais , Linhagem Celular Tumoral , Sobrevivência Celular , Feminino , Regulação Neoplásica da Expressão Gênica , Estimativa de Kaplan-Meier , Luciferases/genética , Camundongos , Transplante de Neoplasias , Proteômica/métodos , Microambiente TumoralRESUMO
BACKGROUND: Checkpoint inhibition has demonstrated clinical efficacy in a variety of solid tumors. Reports of programmed death ligand 1 (PD-L1) expression in glioblastoma are highly variable (ranging from 6% to 88%) and its role as a prognostic marker has yielded conflicting results. OBJECTIVE: To validate the prevalence and prognostic role of PD-L1 expression in a large cohort of diffuse gliomas according to the 2016 revised WHO classification. METHODS: Using tissue microarrays, we compared 5 PD-L1 monoclonal antibodies (n = 56) and validated expression (n = 183) using quantitative immunohistochemistry (IHC) and RNA in situ hybridization (RISH). Expression data from The Cancer Genome Atlas (TCGA) and published studies were compared with clinical outcome. Multiplexed immunophenotyping was used to identify PD-L1+ cell populations in post-treatment glioblastoma. RESULTS: Using a 5% cut-off, PD-L1 expression was significantly associated with a poor prognosis in both histologically defined (n = 125, log-rank P < .001) and recurrent isocitrate dehydrogenase (IDH)-wildtype glioblastoma (n = 60, log-rank P = .015). PD-L1 remained a significant negative prognosticator in Cox regression analysis (hazard ratio: 1.96, P = .021). Analysis of TCGA data confirmed decreased overall survival in recurrent non-glioma CpG island methylator phenotype (G-CIMP) glioblastoma (n = 12, log-rank P = .023), but not in glioblastoma as a group (n = 444, log-rank P = .135). PD-L1 RISH showed a significant correlation with IHC (P < .0001). PD-L1 was observed in the proliferating perivascular stem cell and immune niche of post-treatment glioblastoma. CONCLUSION: A 5% PD-L1 expression cut-off identified a subset of glioblastoma that is associated with a worse clinical outcome. This association remained significant within the newly defined IDH-wildtype classification. These findings could have implications for patient stratification in future clinical trials of PD-1/PD-L1 blockade.
Assuntos
Antígeno B7-H1/metabolismo , Biomarcadores Tumorais/análise , Neoplasias Encefálicas/patologia , Glioblastoma/patologia , Adolescente , Adulto , Antígeno B7-H1/análise , Neoplasias Encefálicas/genética , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Glioblastoma/genética , Humanos , Isocitrato Desidrogenase/genética , Masculino , Pessoa de Meia-Idade , Prognóstico , Modelos de Riscos Proporcionais , Estudos Retrospectivos , Adulto JovemRESUMO
BACKGROUND: Glioblastoma is the most common primary malignancy of the brain, with a dismal prognosis. Immunomodulation via checkpoint inhibition has provided encouraging results in non-CNS malignancies, but prediction of responders has proven to be challenging in glioblastoma patients. OBJECTIVE: To determine the proportion of patients who have a measurable increase of interferon gamma levels in brain tumor tissue after their first dose of nivolumab, and to evaluate the safety of using brain tumor microdialysis to monitor for immune response while evaluating the safety of the combination of anti-programmed death 1 (PD-1) and anti-lymphocyte activation gene 3 (LAG-3) checkpoint inhibition. METHODS: The study design is a single-center, nonrandomized phase 1 clinical trial. Up to 15 adult patients with recurrent glioblastoma will be enrolled with the goal of 10 patients completing the trial over an anticipated 18 mo. Patients will undergo biopsy; placement of microdialysis catheters and lumbar drains; treatment with anti-PD-1 checkpoint inhibition; comprehensive immune biomarker collection; tumor resection; and then treatment with anti-PD-1 and anti-LAG-3 checkpoint inhibition until progression. EXPECTED OUTCOMES: We expect interferon gamma levels to increase in the brain as measured via microdialysis in treated patients. Based on published reports, microdialysis in this patient population is expected to be safe, and anti-LAG-3 and anti-PD-1 combined will likely have a similar side effect profile to other checkpoint inhibitor combinations. DISCUSSION: The failure of recent trials of immune therapies in glioblastoma underscores the need to appropriately measure response in the treated tissue. This trial may provide insight on indicators of which patients will respond to immune therapy.
Assuntos
Neoplasias Encefálicas , Citocinas , Glioblastoma , Microdiálise , Monitorização Imunológica , Adulto , Química Encefálica , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/terapia , Citocinas/análise , Citocinas/isolamento & purificação , Glioblastoma/imunologia , Glioblastoma/metabolismo , Glioblastoma/terapia , Humanos , Interferon gama/análise , Interferon gama/isolamento & purificação , Recidiva Local de Neoplasia/imunologia , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/terapiaRESUMO
Glioblastoma is in need of innovative treatment approaches. Immune therapy for cancer refers to the use of the body's immune system to target malignant cells in the body. Such immune therapeutics have recently been very successful in treating a diverse group of cancerous lesions. As a result, many new immune therapies have gained Food and Drug Administration approval for the treatment of cancer, and there has been an explosion in the study of immune therapeutics for cancer treatment over the past few years. However, the immune suppression of glioblastoma and the unique immune microenvironment of the brain make immune therapeutics more challenging to apply to the brain than to other systemic cancers. Here, we discuss the existing barriers to successful immune therapy for glioblastoma and the ongoing development of immune therapeutics. We will discuss the discovery and classification of immune suppressive factors in the glioblastoma microenvironment; the development of vaccine-based therapies; the use of convection-enhanced delivery to introduce tumoricidal viruses into the tumor microenvironment, leading to secondary immune responses; the emerging use of adoptive cell therapy in the treatment of glioblastoma; and future frontiers, such as the use of cerebral microdialysis for immune monitoring and the use of sequencing to develop patient-specific therapeutics. Armed with a better understanding of the challenges inherent in immune therapy for glioblastoma, we may soon see more successes in immune-based clinical trials for this deadly disease.
RESUMO
Tumor-associated macrophages (TAMs) are predominantly M2 phenotype in solid cancers including hepatocellular carcinoma (HCC). Though differentiation of M2 macrophages has been recently linked to fatty acid oxidation (FAO), whether FAO plays a role in functional maintenance of M2 macrophages is still unclear. Here, we used an in vitro model to mimic TAM-HCC interaction in tumor microenvironment. We found that M2 monocyte-derived macrophages (MDMs) enhanced the proliferation, migration, and invasion of HCC cells through an FAO-dependent way. Further investigations identified that IL-1ß mediated the pro-migratory effect of M2 MDM. Using etomoxir and siRNA to inhibit FAO and palmitate to enhance FAO, we showed that FAO was responsible for the up-regulated secretion of IL-1ß and, thus, the pro-migratory effect in M2 MDMs. In addition, we proved that IL-1ß induction was reactive oxygen species and NLRP3-dependent. Our study demonstrates that FAO plays a key role in functional human M2 macrophages by enhancing IL-1ß secretion to promote HCC cell migration. These findings provide evidence for different dependency of energy sources in macrophages with distinct phenotypes and functions, and suggest a novel strategy to treat HCC by reprogramming cell metabolism or modulating tumor microenvironment.
Assuntos
Carcinoma Hepatocelular/patologia , Movimento Celular/fisiologia , Ácidos Graxos/metabolismo , Interleucina-1beta/metabolismo , Neoplasias Hepáticas/patologia , Macrófagos/fisiologia , Células Cultivadas , Células Hep G2 , Humanos , Macrófagos/citologia , Macrófagos/metabolismo , Oxirredução , Microambiente TumoralRESUMO
We hypothesized that expression of mutant Huntingtin (HTT) would modulate the neurotoxicity of the commonly used organophosphate insecticide, chlorpyrifos (CPF), revealing cellular mechanisms underlying neurodegeneration. Using a mouse striatal cell model of HD, we report that mutant HD cells are more susceptible to CPF-induced cytotoxicity as compared to wild-type. This CPF-induced cytotoxicity caused increased production of reactive oxygen species, reduced glutathione levels, decreased superoxide dismutase activity, and increased malondialdehyde levels in mutant HD cells relative to wild-type. Furthermore, we show that co-treatment with antioxidant agents attenuated the CPF-induced ROS levels and cytotoxicity. Co-treatment with a NADPH oxidase (NOX) inhibitor, apocynin, also attenuated the CPF-induced ROS production and neurotoxicity. CPF caused increased NOX activity in mutant HD lines that was ameliorated following co-treatment with apocynin. Finally, CPF-induced neurotoxicity significantly increased the protein expression of nuclear factor erythroid 2-related factor (Nrf2) in mutant HD cells as compared to wild-type. This study is the first report of CPF-induced toxicity in HD pathophysiology and suggests that mutant HTT and CPF exhibit a disease-toxicant interaction wherein expression of mutant HTT enhances CPF-induced neurotoxicity via a NOX-mediated oxidative stress mechanism to cause neuronal loss in the full length HTT expressing striatal cells.