RESUMO
SARS-CoV-2 infects less than 1% of cells in the human body, yet it can cause severe damage in a variety of organs. Thus, deciphering the non-cell-autonomous effects of SARS-CoV-2 infection is imperative for understanding the cellular and molecular disruption it elicits. Neurological and cognitive defects are among the least understood symptoms of COVID-19 patients, with olfactory dysfunction being their most common sensory deficit. Here, we show that both in humans and hamsters, SARS-CoV-2 infection causes widespread downregulation of olfactory receptors (ORs) and of their signaling components. This non-cell-autonomous effect is preceded by a dramatic reorganization of the neuronal nuclear architecture, which results in dissipation of genomic compartments harboring OR genes. Our data provide a potential mechanism by which SARS-CoV-2 infection alters the cellular morphology and the transcriptome of cells it cannot infect, offering insight to its systemic effects in olfaction and beyond.
Assuntos
Anosmia , COVID-19 , Animais , Cricetinae , Regulação para Baixo , Humanos , Receptores Odorantes , SARS-CoV-2 , OlfatoRESUMO
Melanoma brain metastasis (MBM) frequently occurs in patients with advanced melanoma; yet, our understanding of the underlying salient biology is rudimentary. Here, we performed single-cell/nucleus RNA-seq in 22 treatment-naive MBMs and 10 extracranial melanoma metastases (ECMs) and matched spatial single-cell transcriptomics and T cell receptor (TCR)-seq. Cancer cells from MBM were more chromosomally unstable, adopted a neuronal-like cell state, and enriched for spatially variably expressed metabolic pathways. Key observations were validated in independent patient cohorts, patient-derived MBM/ECM xenograft models, RNA/ATAC-seq, proteomics, and multiplexed imaging. Integrated spatial analyses revealed distinct geography of putative cancer immune evasion and evidence for more abundant intra-tumoral B to plasma cell differentiation in lymphoid aggregates in MBM. MBM harbored larger fractions of monocyte-derived macrophages and dysfunctional TOX+CD8+ T cells with distinct expression of immune checkpoints. This work provides comprehensive insights into MBM biology and serves as a foundational resource for further discovery and therapeutic exploration.
Assuntos
Neoplasias Encefálicas , Melanoma , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/secundário , Linfócitos T CD8-Positivos/patologia , Ecossistema , Humanos , RNA-SeqRESUMO
Lactate accumulates to a significant amount in glioblastomas (GBMs), the most common primary malignant brain tumor with an unfavorable prognosis. However, it remains unclear whether lactate is metabolized by GBMs. Here, we demonstrated that lactate rescued patient-derived xenograft (PDX) GBM cells from nutrient-deprivation-mediated cell death. Transcriptome analysis, ATAC-seq, and ChIP-seq showed that lactate entertained a signature of oxidative energy metabolism. LC/MS analysis demonstrated that U-13C-lactate elicited substantial labeling of TCA-cycle metabolites, acetyl-CoA, and histone protein acetyl-residues in GBM cells. Lactate enhanced chromatin accessibility and histone acetylation in a manner dependent on oxidative energy metabolism and the ATP-citrate lyase (ACLY). Utilizing orthotopic PDX models of GBM, a combined tracer experiment unraveled that lactate carbons were substantially labeling the TCA-cycle metabolites. Finally, pharmacological blockage of oxidative energy metabolism extended overall survival in two orthotopic PDX models in mice. These results establish lactate metabolism as a novel druggable pathway for GBM.
Assuntos
Glioblastoma , Acetilação , Animais , Linhagem Celular Tumoral , Epigênese Genética , Glioblastoma/genética , Glioblastoma/patologia , Histonas/metabolismo , Humanos , Ácido Láctico/metabolismo , CamundongosRESUMO
Alzheimer's disease (AD) is the most common cause of dementia, and disease mechanisms are still not fully understood. Here, we explored pathological changes in human induced pluripotent stem cell (iPSC)-derived neurons carrying the familial AD APPV717I mutation after cell injection into the mouse forebrain. APPV717I mutant iPSCs and isogenic controls were differentiated into neurons revealing enhanced Aß42 production, elevated phospho-tau, and impaired neurite outgrowth in APPV717I neurons. Two months after transplantation, APPV717I and control neural cells showed robust engraftment but at 12 months post-injection, APPV717I grafts were smaller and demonstrated impaired neurite outgrowth compared to controls, while plaque and tangle pathology were not seen. Single-nucleus RNA-sequencing of micro-dissected grafts, performed 2 months after cell injection, identified significantly altered transcriptome signatures in APPV717I iPSC-derived neurons pointing towards dysregulated synaptic function and axon guidance. Interestingly, APPV717I neurons showed an increased expression of genes, many of which are also upregulated in postmortem neurons of AD patients including the transmembrane protein LINGO2. Downregulation of LINGO2 in cultured APPV717I neurons rescued neurite outgrowth deficits and reversed key AD-associated transcriptional changes related but not limited to synaptic function, apoptosis and cellular senescence. These results provide important insights into transcriptional dysregulation in xenografted APPV717I neurons linked to synaptic function, and they indicate that LINGO2 may represent a potential therapeutic target in AD.
Assuntos
Doença de Alzheimer , Precursor de Proteína beta-Amiloide , Células-Tronco Pluripotentes Induzidas , Neurônios , Transcriptoma , Humanos , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Doença de Alzheimer/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Animais , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Mutação , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Sinapses/patologia , Sinapses/metabolismo , Peptídeos beta-Amiloides/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologiaRESUMO
Infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is associated with acute and postacute cognitive and neuropsychiatric symptoms including impaired memory, concentration, attention, sleep and affect. Mechanisms underlying these brain symptoms remain understudied. Here we report that SARS-CoV-2-infected hamsters exhibit a lack of viral neuroinvasion despite aberrant blood-brain barrier permeability. Hamsters and patients deceased from coronavirus disease 2019 (COVID-19) also exhibit microglial activation and expression of interleukin (IL)-1ß and IL-6, especially within the hippocampus and the medulla oblongata, when compared with non-COVID control hamsters and humans who died from other infections, cardiovascular disease, uraemia or trauma. In the hippocampal dentate gyrus of both COVID-19 hamsters and humans, we observed fewer neuroblasts and immature neurons. Protracted inflammation, blood-brain barrier disruption and microglia activation may result in altered neurotransmission, neurogenesis and neuronal damage, explaining neuropsychiatric presentations of COVID-19. The involvement of the hippocampus may explain learning, memory and executive dysfunctions in COVID-19 patients.
Assuntos
COVID-19 , Humanos , Citocinas , SARS-CoV-2 , Hipocampo , Neurogênese/fisiologiaRESUMO
While several studies have attributed the development of tumour-associated seizures to an excitatory-inhibitory imbalance, we have yet to resolve the spatiotemporal interplay between different types of neuron in glioma-infiltrated cortex. Herein, we combined methods for single unit analysis of microelectrode array recordings with wide-field optical mapping of Thy1-GCaMP pyramidal cells in an ex vivo acute slice model of diffusely infiltrating glioma. This enabled simultaneous tracking of individual neurons from both excitatory and inhibitory populations throughout seizure-like events. Moreover, our approach allowed for observation of how the crosstalk between these neurons varied spatially, as we recorded across an extended region of glioma-infiltrated cortex. In tumour-bearing slices, we observed marked alterations in single units classified as putative fast-spiking interneurons, including reduced firing, activity concentrated within excitatory bursts and deficits in local inhibition. These results were correlated with increases in overall excitability. Mechanistic perturbation of this system with the mTOR inhibitor AZD8055 revealed increased firing of putative fast-spiking interneurons and restoration of local inhibition, with concomitant decreases in overall excitability. Altogether, our findings suggest that diffusely infiltrating glioma affect the interplay between excitatory and inhibitory neuronal populations in a reversible manner, highlighting a prominent role for functional mechanisms linked to mTOR activation.
Assuntos
Glioma , Células Piramidais , Humanos , Potenciais de Ação/fisiologia , Células Piramidais/fisiologia , Neurônios/fisiologia , Convulsões , Serina-Treonina Quinases TORRESUMO
Post-zygotically acquired genetic variants, or somatic variants, that arise during cortical development have emerged as important causes of focal epilepsies, particularly those due to malformations of cortical development. Pathogenic somatic variants have been identified in many genes within the PI3K-AKT-mTOR-signalling pathway in individuals with hemimegalencephaly and focal cortical dysplasia (type II), and more recently in SLC35A2 in individuals with focal cortical dysplasia (type I) or non-dysplastic epileptic cortex. Given the expanding role of somatic variants across different brain malformations, we sought to delineate the landscape of somatic variants in a large cohort of patients who underwent epilepsy surgery with hemimegalencephaly or focal cortical dysplasia. We evaluated samples from 123 children with hemimegalencephaly (n = 16), focal cortical dysplasia type I and related phenotypes (n = 48), focal cortical dysplasia type II (n = 44), or focal cortical dysplasia type III (n = 15). We performed high-depth exome sequencing in brain tissue-derived DNA from each case and identified somatic single nucleotide, indel and large copy number variants. In 75% of individuals with hemimegalencephaly and 29% with focal cortical dysplasia type II, we identified pathogenic variants in PI3K-AKT-mTOR pathway genes. Four of 48 cases with focal cortical dysplasia type I (8%) had a likely pathogenic variant in SLC35A2. While no other gene had multiple disease-causing somatic variants across the focal cortical dysplasia type I cohort, four individuals in this group had a single pathogenic or likely pathogenic somatic variant in CASK, KRAS, NF1 and NIPBL, genes previously associated with neurodevelopmental disorders. No rare pathogenic or likely pathogenic somatic variants in any neurological disease genes like those identified in the focal cortical dysplasia type I cohort were found in 63 neurologically normal controls (P = 0.017), suggesting a role for these novel variants. We also identified a somatic loss-of-function variant in the known epilepsy gene, PCDH19, present in a small number of alleles in the dysplastic tissue from a female patient with focal cortical dysplasia IIIa with hippocampal sclerosis. In contrast to focal cortical dysplasia type II, neither focal cortical dysplasia type I nor III had somatic variants in genes that converge on a unifying biological pathway, suggesting greater genetic heterogeneity compared to type II. Importantly, we demonstrate that focal cortical dysplasia types I, II and III are associated with somatic gene variants across a broad range of genes, many associated with epilepsy in clinical syndromes caused by germline variants, as well as including some not previously associated with radiographically evident cortical brain malformations.
Assuntos
Epilepsia , Hemimegalencefalia , Malformações do Desenvolvimento Cortical , Caderinas , Proteínas de Ciclo Celular , Feminino , Humanos , Malformações do Desenvolvimento Cortical do Grupo I , Mutação , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Protocaderinas , Serina-Treonina Quinases TORRESUMO
BACKGROUND: Topotecan is cytotoxic to glioma cells but is clinically ineffective because of drug delivery limitations. Systemic delivery is limited by toxicity and insufficient brain penetrance, and, to date, convection-enhanced delivery (CED) has been restricted to a single treatment of restricted duration. To address this problem, we engineered a subcutaneously implanted catheter-pump system capable of repeated, chronic (prolonged, pulsatile) CED of topotecan into the brain and tested its safety and biological effects in patients with recurrent glioblastoma. METHODS: We did a single-centre, open-label, single-arm, phase 1b clinical trial at Columbia University Irving Medical Center (New York, NY, USA). Eligible patients were at least 18 years of age with solitary, histologically confirmed recurrent glioblastoma showing radiographic progression after surgery, radiotherapy, and chemotherapy, and a Karnofsky Performance Status of at least 70. Five patients had catheters stereotactically implanted into the glioma-infiltrated peritumoural brain and connected to subcutaneously implanted pumps that infused 146 µM topotecan 200 µL/h for 48 h, followed by a 5-7-day washout period before the next infusion, with four total infusions. After the fourth infusion, the pump was removed and the tumour was resected. The primary endpoint of the study was safety of the treatment regimen as defined by presence of serious adverse events. Analyses were done in all treated patients. The trial is closed, and is registered with ClinicalTrials.gov, NCT03154996. FINDINGS: Between Jan 22, 2018, and July 8, 2019, chronic CED of topotecan was successfully completed safely in all five patients, and was well tolerated without substantial complications. The only grade 3 adverse event related to treatment was intraoperative supplemental motor area syndrome (one [20%] of five patients in the treatment group), and there were no grade 4 adverse events. Other serious adverse events were related to surgical resection and not the study treatment. Median follow-up was 12 months (IQR 10-17) from pump explant. Post-treatment tissue analysis showed that topotecan significantly reduced proliferating tumour cells in all five patients. INTERPRETATION: In this small patient cohort, we showed that chronic CED of topotecan is a potentially safe and active therapy for recurrent glioblastoma. Our analysis provided a unique tissue-based assessment of treatment response without the need for large patient numbers. This novel delivery of topotecan overcomes limitations in delivery and treatment response assessment for patients with glioblastoma and could be applicable for other anti-glioma drugs or other CNS diseases. Further studies are warranted to determine the effect of this drug delivery approach on clinical outcomes. FUNDING: US National Institutes of Health, The William Rhodes and Louise Tilzer Rhodes Center for Glioblastoma, the Michael Weiner Glioblastoma Research Into Treatment Fund, the Gary and Yael Fegel Foundation, and The Khatib Foundation.
Assuntos
Glioblastoma , Glioma , Humanos , Topotecan/efeitos adversos , Glioblastoma/tratamento farmacológico , Convecção , Recidiva Local de Neoplasia/tratamento farmacológico , Glioma/patologiaRESUMO
Understanding dexamethasone's effect on the immune microenvironment in glioma patients is of key importance. We performed a comprehensive literature review using the NCBI PubMed database for all articles meeting the following search criteria. ((dexamethasone[All Fields]) AND (glioma or glioblastoma)[Title/Abstract]) AND (immune or T cell or B cell or monocyte or neutrophil or macrophage). Forty-three manuscripts were deemed relevant to the topic at hand. Multiple clinical studies have linked dexamethasone use to decreased overall survival while preclinical studies in murine glioma models have demonstrated decreased tumor-infiltrating lymphocytes after dexamethasone administration.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Glioma , Animais , Neoplasias Encefálicas/tratamento farmacológico , Dexametasona/uso terapêutico , Glioma/tratamento farmacológico , Humanos , Imunoterapia , Linfócitos do Interstício Tumoral , Camundongos , Medicina de Precisão , Microambiente TumoralRESUMO
Many patients with SARS-CoV-2 infection develop neurological signs and symptoms; although, to date, little evidence exists that primary infection of the brain is a significant contributing factor. We present the clinical, neuropathological and molecular findings of 41 consecutive patients with SARS-CoV-2 infections who died and underwent autopsy in our medical centre. The mean age was 74 years (38-97 years), 27 patients (66%) were male and 34 (83%) were of Hispanic/Latinx ethnicity. Twenty-four patients (59%) were admitted to the intensive care unit. Hospital-associated complications were common, including eight patients (20%) with deep vein thrombosis/pulmonary embolism, seven (17%) with acute kidney injury requiring dialysis and 10 (24%) with positive blood cultures during admission. Eight (20%) patients died within 24 h of hospital admission, while 11 (27%) died more than 4 weeks after hospital admission. Neuropathological examination of 20-30 areas from each brain revealed hypoxic/ischaemic changes in all brains, both global and focal; large and small infarcts, many of which appeared haemorrhagic; and microglial activation with microglial nodules accompanied by neuronophagia, most prominently in the brainstem. We observed sparse T lymphocyte accumulation in either perivascular regions or in the brain parenchyma. Many brains contained atherosclerosis of large arteries and arteriolosclerosis, although none showed evidence of vasculitis. Eighteen patients (44%) exhibited pathologies of neurodegenerative diseases, which was not unexpected given the age range of our patients. We examined multiple fresh frozen and fixed tissues from 28 brains for the presence of viral RNA and protein, using quantitative reverse-transcriptase PCR, RNAscope® and immunocytochemistry with primers, probes and antibodies directed against the spike and nucleocapsid regions. The PCR analysis revealed low to very low, but detectable, viral RNA levels in the majority of brains, although they were far lower than those in the nasal epithelia. RNAscope® and immunocytochemistry failed to detect viral RNA or protein in brains. Our findings indicate that the levels of detectable virus in coronavirus disease 2019 brains are very low and do not correlate with the histopathological alterations. These findings suggest that microglial activation, microglial nodules and neuronophagia, observed in the majority of brains, do not result from direct viral infection of brain parenchyma, but more likely from systemic inflammation, perhaps with synergistic contribution from hypoxia/ischaemia. Further studies are needed to define whether these pathologies, if present in patients who survive coronavirus disease 2019, might contribute to chronic neurological problems.
Assuntos
Infarto Encefálico/patologia , Encéfalo/patologia , COVID-19/patologia , Hipóxia-Isquemia Encefálica/patologia , Hemorragias Intracranianas/patologia , Injúria Renal Aguda/complicações , Injúria Renal Aguda/fisiopatologia , Injúria Renal Aguda/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Bacteriemia/complicações , Encéfalo/metabolismo , Infarto Encefálico/complicações , COVID-19/complicações , COVID-19/fisiopatologia , Proteínas do Nucleocapsídeo de Coronavírus/metabolismo , Feminino , Humanos , Hipóxia-Isquemia Encefálica/complicações , Inflamação , Unidades de Terapia Intensiva , Hemorragias Intracranianas/complicações , Masculino , Microglia/patologia , Pessoa de Meia-Idade , Neurônios/patologia , Fagocitose , Fosfoproteínas/metabolismo , Embolia Pulmonar/complicações , Embolia Pulmonar/fisiopatologia , RNA Viral/metabolismo , Diálise Renal , Reação em Cadeia da Polimerase Via Transcriptase Reversa , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/metabolismo , Taxa de Sobrevida , Linfócitos T/patologia , Trombose Venosa/complicações , Trombose Venosa/fisiopatologiaRESUMO
The ability of glioblastoma to disperse through the brain contributes to its lethality, and blocking this behavior has been an appealing therapeutic approach. Although a number of proinvasive signaling pathways are active in glioblastoma, many are redundant, so targeting one can be overcome by activating another. However, these pathways converge on nonredundant components of the cytoskeleton, and we have shown that inhibiting one of these-the myosin II family of cytoskeletal motors-blocks glioblastoma invasion even with simultaneous activation of multiple upstream promigratory pathways. Myosin IIA and IIB are the most prevalent isoforms of myosin II in glioblastoma, and we now show that codeleting these myosins markedly impairs tumorigenesis and significantly prolongs survival in a rodent model of this disease. However, while targeting just myosin IIA also impairs tumor invasion, it surprisingly increases tumor proliferation in a manner that depends on environmental mechanics. On soft surfaces myosin IIA deletion enhances ERK1/2 activity, while on stiff surfaces it enhances the activity of NFκB, not only in glioblastoma but in triple-negative breast carcinoma and normal keratinocytes as well. We conclude myosin IIA suppresses tumorigenesis in at least two ways that are modulated by the mechanics of the tumor and its stroma. Our results also suggest that inhibiting tumor invasion can enhance tumor proliferation and that effective therapy requires targeting cellular components that drive both proliferation and invasion simultaneously.
Assuntos
Carcinogênese/metabolismo , Citoesqueleto/metabolismo , Glioblastoma/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas de Neoplasias/metabolismo , Miosina não Muscular Tipo IIA/metabolismo , Animais , Carcinogênese/genética , Carcinogênese/patologia , Linhagem Celular Tumoral , Citoesqueleto/genética , Citoesqueleto/patologia , Glioblastoma/genética , Glioblastoma/patologia , Camundongos , Proteínas de Neoplasias/genética , Miosina não Muscular Tipo IIA/genéticaRESUMO
Glioblastomas are aggressive primary brain tumors known for their inter- and intratumor heterogeneity. This disease is uniformly fatal, with intratumor heterogeneity the major reason for treatment failure and recurrence. Just like the nature vs nurture debate, heterogeneity can arise from intrinsic or environmental influences. Whilst it is impossible to clinically separate observed behavior of cells from their environmental context, using a mathematical framework combined with multiscale data gives us insight into the relative roles of variation from different sources. To better understand the implications of intratumor heterogeneity on therapeutic outcomes, we created a hybrid agent-based mathematical model that captures both the overall tumor kinetics and the individual cellular behavior. We track single cells as agents, cell density on a coarser scale, and growth factor diffusion and dynamics on a finer scale over time and space. Our model parameters were fit utilizing serial MRI imaging and cell tracking data from ex vivo tissue slices acquired from a growth-factor driven glioblastoma murine model. When fitting our model to serial imaging only, there was a spectrum of equally-good parameter fits corresponding to a wide range of phenotypic behaviors. When fitting our model using imaging and cell scale data, we determined that environmental heterogeneity alone is insufficient to match the single cell data, and intrinsic heterogeneity is required to fully capture the migration behavior. The wide spectrum of in silico tumors also had a wide variety of responses to an application of an anti-proliferative treatment. Recurrent tumors were generally less proliferative than pre-treatment tumors as measured via the model simulations and validated from human GBM patient histology. Further, we found that all tumors continued to grow with an anti-migratory treatment alone, but the anti-proliferative/anti-migratory combination generally showed improvement over an anti-proliferative treatment alone. Together our results emphasize the need to better understand the underlying phenotypes and tumor heterogeneity present in a tumor when designing therapeutic regimens.
Assuntos
Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/fisiopatologia , Glioblastoma/diagnóstico por imagem , Glioblastoma/fisiopatologia , Imageamento por Ressonância Magnética , Animais , Proliferação de Células , Biologia Computacional , Simulação por Computador , Humanos , Cinética , Masculino , Camundongos Endogâmicos NOD , Modelos Teóricos , Fenótipo , Ratos , Ratos Sprague-DawleyRESUMO
Intramedullary spinal cord tumors are a rare and understudied cancer with poor treatment options and prognosis. Our prior study used a combination of PDGF-B, HRAS, and p53 knockdown to induce the development of high-grade glioma in the spinal cords of minipigs. In this study, we evaluate the ability of each vector alone and combinations of vectors to produce high-grade spinal cord gliomas. Eight groups of rats (n = 8/group) underwent thoracolumbar laminectomy and injection of lentiviral vector in the lateral white matter of the spinal cord. Each group received a different combination of lentiviral vectors expressing PDGF-B, a constitutively active HRAS mutant, or shRNA targeting p53, or a control vector. All animals were monitored once per week for clinical deficits for 98 days. Tissues were harvested and analyzed using hematoxylin and eosin (H&E) and immunohistochemical (IHC) staining. Rats injected with PDGF-B+HRAS+sh-p53 (triple cocktail) exhibited statistically significant declines in all behavioral measures (Basso Beattie Bresnahan scoring, Tarlov scoring, weight, and survival rate) over time when compared to the control. Histologically, all groups except the control and those injected with sh-p53 displayed the development of tumors at the injection site, although there were differences in the rate of tumor growth and the histopathological features of the lesions between groups. Examination of immunohistochemistry revealed rats receiving triple cocktail displayed the largest and most significant increase in the Ki67 proliferation index and GFAP positivity than any other group. PDGF-B+HRAS also displayed a significant increase in the Ki67 proliferation index. Rats receiving PDGF-B alone and PDGF-B+ sh-p53 displayed more a significant increase in SOX2-positive staining than in any other group. We found that different vector combinations produced differing high-grade glioma models in rodents. The combination of all three vectors produced a model of high-grade glioma more efficiently and aggressively with respect to behavioral, physiological, and histological characteristics than the rest of the vector combinations. Thus, the present rat model of spinal cord glioma may potentially be used to evaluate therapeutic strategies in the future.
Assuntos
Glioma/etiologia , Lentivirus/genética , Neoplasias da Medula Espinal/etiologia , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Proliferação de Células , Vetores Genéticos , Glioma/patologia , Glioma/fisiopatologia , Mutação , Neoplasias Experimentais/etiologia , Neoplasias Experimentais/patologia , Neoplasias Experimentais/fisiopatologia , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/metabolismo , Fator de Crescimento Derivado de Plaquetas/genética , Fator de Crescimento Derivado de Plaquetas/metabolismo , RNA Interferente Pequeno/administração & dosagem , RNA Interferente Pequeno/genética , Ratos , Ratos Sprague-Dawley , Neoplasias da Medula Espinal/patologia , Neoplasias da Medula Espinal/fisiopatologia , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas ras/genética , Proteínas ras/metabolismoRESUMO
Glioblastoma (GB) is one of the deadliest brain cancers to afflict humans, and it has a very poor survival rate even with treatment. The extracellular adenosine-generating enzyme CD73 is involved in many cellular functions that can be usurped by tumors, including cell adhesion, proliferation, invasion, and angiogenesis. We set out to determine the role of CD73 in GB pathogenesis. To do this, we established a unique GB mouse model (CD73-FLK) in which we spatially expressed CD73 on endothelial cells in CD73-/- mice. This allowed us to elucidate the mechanism of host CD73 versus GB-expressed CD73 by comparing GB pathogenesis in WT, CD73-/-, and CD73-FLK mice. GB in CD73-/- mice had decreased tumor size, decreased tumor vessel density, and reduced tumor invasiveness compared with GB in WT mice. Interestingly, GBs in CD73-FLK mice were much more invasive and caused complete distortion of the brain morphology. We showed a 20-fold upregulation of A2B AR on GB compared with sham, and its activation induced matrix metalloproteinase-2, which enhanced GB pathogenesis. Inhibition of A2B AR signaling decreased multidrug resistance transporter protein expression, including permeability glycoprotein (P-gp) and multidrug resistance-associated protein 1 (MRP1). Further, we showed that blockade of A2B AR signaling potently increased GB cell death induced by the chemotherapeutic drug temozolomide. Together, these findings suggest that CD73 and A2B AR play a multifaceted role in GB pathogenesis and progression and that targeting the CD73-A2B AR axis can benefit GB patients and inform new approaches for therapy to treat GB patients.SIGNIFICANCE STATEMENT Glioblastoma (GB) is the most devastating primary brain tumor. GB patients' median survival is 16 months even with treatment. It is critical that we develop prophylaxes to advance GB treatment and improve patient survival. CD73-generated adenosine has been implicated in cancer pathogenesis, but its role in GB was not ascertained. Here, we demonstrated that host CD73 plays a prominent role in multiple areas of glioblastoma pathogenesis, including promoting GB growth, its angiogenesis, and its invasiveness. We found a 20-fold increase in A2B adenosine receptor (AR) expression on GB compared with sham, and its inhibition increased GB chemosensitivity to temozolomide. These findings strongly indicate that blockade or inhibition of CD73 and the A2B AR are prime targets for future GB therapy.
Assuntos
5'-Nucleotidase/metabolismo , Neoplasias Encefálicas/metabolismo , Resistencia a Medicamentos Antineoplásicos/fisiologia , Glioblastoma/metabolismo , Receptor A2B de Adenosina/metabolismo , Animais , Neoplasias Encefálicas/patologia , Glioblastoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/fisiologiaRESUMO
The purpose of this study is to develop a platform in which the cellular and molecular underpinnings of chronic focal neocortical lesional epilepsy can be explored and use it to characterize seizure-like events (SLEs) in an ex vivo model of infiltrating high-grade glioma. Microelectrode arrays were used to study electrophysiologic changes in ex vivo acute brain slices from a PTEN/p53 deleted, PDGF-B driven mouse model of high-grade glioma. Electrode locations were co-registered to the underlying histology to ascertain the influence of the varying histologic landscape on the observed electrophysiologic changes. Peritumoral, infiltrated, and tumor sites were sampled in tumor-bearing slices. Following the addition of zero Mg2+ solution, all three histologic regions in tumor-bearing slices showed significantly greater increases in firing rates when compared to the control sites. Tumor-bearing slices demonstrated increased proclivity for SLEs, with 40 events in tumor-bearing slices and 5 events in control slices (p-valueâ¯=â¯.0105). Observed SLEs were characterized by either low voltage fast (LVF) onset patterns or short bursts of repetitive widespread, high amplitude low frequency discharges. Seizure foci comprised areas from all three histologic regions. The onset electrode was found to be at the infiltrated margin in 50% of cases and in the peritumoral region in 36.9% of cases. These findings reveal a landscape of histopathologic and electrophysiologic alterations associated with ictogenesis and spread of tumor-associated seizures.
Assuntos
Neoplasias Encefálicas/fisiopatologia , Encéfalo/fisiopatologia , Glioma/fisiopatologia , Neurônios/fisiologia , Convulsões/fisiopatologia , Potenciais de Ação , Animais , Neoplasias Encefálicas/complicações , Modelos Animais de Doenças , Glioma/complicações , Camundongos Transgênicos , Microeletrodos , Convulsões/complicaçõesRESUMO
Common approaches to gene signature discovery in single-cell RNA-sequencing (scRNA-seq) depend upon predefined structures like clusters or pseudo-temporal order, require prior normalization, or do not account for the sparsity of single-cell data. We present single-cell hierarchical Poisson factorization (scHPF), a Bayesian factorization method that adapts hierarchical Poisson factorization (Gopalan et al, 2015, Proceedings of the 31st Conference on Uncertainty in Artificial Intelligence, 326) for de novo discovery of both continuous and discrete expression patterns from scRNA-seq. scHPF does not require prior normalization and captures statistical properties of single-cell data better than other methods in benchmark datasets. Applied to scRNA-seq of the core and margin of a high-grade glioma, scHPF uncovers marked differences in the abundance of glioma subpopulations across tumor regions and regionally associated expression biases within glioma subpopulations. scHFP revealed an expression signature that was spatially biased toward the glioma-infiltrated margins and associated with inferior survival in glioblastoma.
Assuntos
Glioma/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Análise de Célula Única , Transcriptoma/genética , Teorema de Bayes , Regulação Neoplásica da Expressão Gênica/genética , Glioma/patologia , Humanos , Distribuição de PoissonRESUMO
BACKGROUND: Sex is recognized as a significant determinant of outcome among glioblastoma patients, but the relative prognostic importance of glioblastoma features has not been thoroughly explored for sex differences. METHODS: Combining multi-modal MR images, biomathematical models, and patient clinical information, this investigation assesses which pretreatment variables have a sex-specific impact on the survival of glioblastoma patients (299 males and 195 females). RESULTS: Among males, tumor (T1Gd) radius was a predictor of overall survival (HR = 1.027, p = 0.044). Among females, higher tumor cell net invasion rate was a significant detriment to overall survival (HR = 1.011, p < 0.001). Female extreme survivors had significantly smaller tumors (T1Gd) (p = 0.010 t-test), but tumor size was not correlated with female overall survival (p = 0.955 CPH). Both male and female extreme survivors had significantly lower tumor cell net proliferation rates than other patients (M p = 0.004, F p = 0.001, t-test). CONCLUSION: Despite similar distributions of the MR imaging parameters between males and females, there was a sex-specific difference in how these parameters related to outcomes.
Assuntos
Neoplasias Encefálicas/mortalidade , Glioblastoma/mortalidade , Imageamento por Ressonância Magnética/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/terapia , Criança , Feminino , Seguimentos , Glioblastoma/patologia , Glioblastoma/terapia , Humanos , Interpretação de Imagem Assistida por Computador , Masculino , Pessoa de Meia-Idade , Modelos Teóricos , Prognóstico , Estudos Retrospectivos , Fatores Sexuais , Taxa de Sobrevida , Adulto JovemRESUMO
OBJECTIVE: Somatic variants are a recognized cause of epilepsy-associated focal malformations of cortical development (MCD). We hypothesized that somatic variants may underlie a wider range of focal epilepsy, including nonlesional focal epilepsy (NLFE). Through genetic analysis of brain tissue, we evaluated the role of somatic variation in focal epilepsy with and without MCD. METHODS: We identified somatic variants through high-depth exome and ultra-high-depth candidate gene sequencing of DNA from epilepsy surgery specimens and leukocytes from 18 individuals with NLFE and 38 with focal MCD. RESULTS: We observed somatic variants in 5 cases in SLC35A2, a gene associated with glycosylation defects and rare X-linked epileptic encephalopathies. Nonsynonymous variants in SLC35A2 were detected in resected brain, and absent from leukocytes, in 3 of 18 individuals (17%) with NLFE, 1 female and 2 males, with variant allele frequencies (VAFs) in brain-derived DNA of 2 to 14%. Pathologic evaluation revealed focal cortical dysplasia type Ia (FCD1a) in 2 of the 3 NLFE cases. In the MCD cohort, nonsynonymous variants in SCL35A2 were detected in the brains of 2 males with intractable epilepsy, developmental delay, and magnetic resonance imaging suggesting FCD, with VAFs of 19 to 53%; Evidence for FCD was not observed in either brain tissue specimen. INTERPRETATION: We report somatic variants in SLC35A2 as an explanation for a substantial fraction of NLFE, a largely unexplained condition, as well as focal MCD, previously shown to result from somatic mutation but until now only in PI3K-AKT-mTOR pathway genes. Collectively, our findings suggest a larger role than previously recognized for glycosylation defects in the intractable epilepsies. Ann Neurol 2018.
Assuntos
Encéfalo/patologia , Epilepsia Resistente a Medicamentos/genética , Proteínas de Transporte de Monossacarídeos/genética , Neocórtex/patologia , Adolescente , Criança , Exoma/genética , Feminino , Humanos , Masculino , Malformações do Desenvolvimento Cortical/genética , Mutação/genética , Neurônios/patologia , Fosfatidilinositol 3-Quinases/genética , Serina-Treonina Quinases TOR/genética , Adulto JovemRESUMO
BACKGROUND: National guidelines recommend maximal safe resection of low-grade and high-grade oligodendrogliomas. However, there is no level 1 evidence to support these guidelines, and recent retrospective studies on the topic have yielded mixed results. OBJECTIVE: To assess the association between extent of resection (EOR) and survival for oligodendrogliomas in the general U.S. METHODS: Cases diagnosed between 2004 and 2013 were selected from the Surveillance, Epidemiology, and End-Results (SEER) Program and retrospectively analyzed for treatment, prognostic factors, and survival times. Cases that did not undergo tumor de-bulking surgery (e.g. no surgery or biopsy alone) were compared to subtotal resection (resection) and gross-total resection (GTR). The primary end-points were overall survival (OS) and cause-specific survival (CSS). An external validation cohort with 1p/19q-codeleted tumors was creating using the TCGA and GSE16011 datasets. RESULTS: 3135 Cases were included in the final analysis. The 75% survival time (75ST) and 5-year survival rates were 47 months and 70.8%, respectively. Subtotal resection (STR, 75ST = 50 months) and GTR (75ST = 61 months) were associated with improved survival times compared to cases that did not undergo surgical debulking (75ST = 20 months, P < 0.001 for both), with reduced hazard ratios (HRs) after controlling for other factors (HR 0.81 [0.68-0.97] and HR 0.65 [0.54-0.79], respectively). GTR was associated with improved OS in both low-grade and anaplastic oligodendroglioma subgroups (HR 0.74 [0.58-0.95], HR 0.60 [0.44-0.82], respectively) while STR fell short of significance in the subgroup analysis. All findings were corroborated by multivariable analysis of CSS and externally validated in a cohort of patients with 1p19q-codeleted tumors. CONCLUSION: Greater EOR is associated with improved survival in oligodendrogliomas. Our findings in this U.S. population-based cohort support national guidelines.
Assuntos
Neoplasias Encefálicas/mortalidade , Procedimentos Neurocirúrgicos/mortalidade , Oligodendroglioma/mortalidade , Adolescente , Adulto , Neoplasias Encefálicas/epidemiologia , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/cirurgia , Criança , Pré-Escolar , Feminino , Seguimentos , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Procedimentos Neurocirúrgicos/métodos , Oligodendroglioma/epidemiologia , Oligodendroglioma/patologia , Oligodendroglioma/cirurgia , Prognóstico , Estudos Retrospectivos , Taxa de Sobrevida , Estados Unidos/epidemiologia , Adulto JovemRESUMO
Paracrine PDGF signaling is involved in many processes in the body, both normal and pathological, including embryonic development, angiogenesis, and wound healing as well as liver fibrosis, atherosclerosis, and cancers. We explored this seemingly dual (normal and pathological) role of PDGF mathematically by modeling the release of PDGF in brain tissue and then varying the dynamics of this release. Resulting simulations show that by varying the dynamics of a PDGF source, our model predicts three possible outcomes for PDGF-driven cellular recruitment and lesion growth: (1) localized, short duration of growth, (2) localized, chronic growth, and (3) widespread chronic growth. Further, our model predicts that the type of response is much more sensitive to the duration of PDGF exposure than the maximum level of that exposure. This suggests that extended duration of paracrine PDGF signal during otherwise normal processes could potentially lead to lesions having a phenotype consistent with pathologic conditions.