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1.
J Neurooncol ; 165(1): 161-169, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37878192

RESUMO

BACKGROUND: Neurocognitive deficits are common in pediatric brain tumor survivors. The use of single nucleotide polymorphism (SNP) analysis in DNA repair genes may identify children treated with radiation therapy for brain tumors at increased risk for treatment toxicity and adverse neurocognitive outcomes. MATERIALS: The Human 660W-Quad v1.0 DNA BeadChip analysis (Illumina) was used to evaluate 1048 SNPs from 59 DNA repair genes in 46 subjects. IQ testing was measured by the Wechsler Intelligence Scale for Children. Linear regression was used to identify the 10 SNPs with the strongest association with IQ scores while adjusting for radiation type. RESULTS: The low vs high IQ patient cohorts were well matched for time from first treatment to most recent IQ, first treatment age, sex, and treatments received. 5 SNPs on 3 different genes (CYP29, XRCC1, and BRCA1) and on 3 different chromosomes (10, 19, and 17) had the strongest association with most recent IQ score that was not modified by radiation type. Furthermore, 5 SNPs on 4 different genes (WRN, NR3C1, ERCC4, RAD51L1) on 4 different chromosomes (8, 5, 16, 14) had the strongest association with change in IQ independent of radiation type, first IQ, and years between IQ measures. CONCLUSIONS: SNPs offer the potential to predict adverse neurocognitive outcomes in pediatric brain tumor survivors. Our results require validation in a larger patient cohort. Improving the ability to identify children at risk of treatment related neurocognitive deficits could allow for better treatment stratification and early cognitive interventions.


Assuntos
Neoplasias Encefálicas , Criança , Humanos , Neoplasias Encefálicas/complicações , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/radioterapia , Testes de Inteligência , Sobreviventes , Irradiação Craniana/efeitos adversos , Testes Neuropsicológicos , Proteína 1 Complementadora Cruzada de Reparo de Raio-X
2.
Acta Neuropathol ; 142(2): 339-360, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34046693

RESUMO

Ependymomas (EPN) are central nervous system tumors comprising both aggressive and more benign molecular subtypes. However, therapy of the high-risk subtypes posterior fossa group A (PF-A) and supratentorial RELA-fusion positive (ST-RELA) is limited to gross total resection and radiotherapy, as effective systemic treatment concepts are still lacking. We have recently described fibroblast growth factor receptors 1 and 3 (FGFR1/FGFR3) as oncogenic drivers of EPN. However, the underlying molecular mechanisms and their potential as therapeutic targets have not yet been investigated in detail. Making use of transcriptomic data across 467 EPN tissues, we found that FGFR1 and FGFR3 were both widely expressed across all molecular groups. FGFR3 mRNA levels were enriched in ST-RELA showing the highest expression among EPN as well as other brain tumors. We further identified high expression levels of fibroblast growth factor 1 and 2 (FGF1, FGF2) across all EPN subtypes while FGF9 was elevated in ST-EPN. Interrogation of our EPN single-cell RNA-sequencing data revealed that FGFR3 was further enriched in cycling and progenitor-like cell populations. Corroboratively, we found FGFR3 to be predominantly expressed in radial glia cells in both mouse embryonal and human brain datasets. Moreover, we detected alternative splicing of the FGFR1/3-IIIc variant, which is known to enhance ligand affinity and FGFR signaling. Dominant-negative interruption of FGFR1/3 activation in PF-A and ST-RELA cell models demonstrated inhibition of key oncogenic pathways leading to reduced cell growth and stem cell characteristics. To explore the feasibility of therapeutically targeting FGFR, we tested a panel of FGFR inhibitors in 12 patient-derived EPN cell models revealing sensitivity in the low-micromolar to nano-molar range. Finally, we gain the first clinical evidence for the activity of the FGFR inhibitor nintedanib in the treatment of a patient with recurrent ST-RELA. Together, these preclinical and clinical data suggest FGFR inhibition as a novel and feasible approach to combat aggressive EPN.


Assuntos
Ependimoma/patologia , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Animais , Neoplasias do Sistema Nervoso Central/genética , Neoplasias do Sistema Nervoso Central/patologia , Ependimoma/genética , Humanos , Camundongos , Recidiva Local de Neoplasia/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/metabolismo , Receptores de Fatores de Crescimento de Fibroblastos/genética
3.
J Neurooncol ; 148(3): 569-575, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32506370

RESUMO

INTRODUCTION: Low-grade glioma (LGG) represent the most common pediatric central nervous system tumor. When total surgical resection is not feasible, chemotherapy is first-line therapy in children. Multiple pediatric LGG chemotherapy regimens have been investigated with variable 2-year event free survival (EFS) rates of 39-69%. To date, treatment of pediatric LGG with a carboplatin and vinblastine (C/VBL) chemotherapy regimen has only been evaluated in a phase 1 dose-finding study. METHODS: A retrospective review of pediatric patients with LGG who were treated with C/VBL at Children's Hospital of Colorado or Akron Children's Hospital from 2011 to 2017 was conducted. Data collected included patient demographics, tumor location, disease response, neurofibromatosis 1 (NF1) status, therapy duration and toxicities. Response to therapy was determined by objective findings on imaging and treating physicians' evaluation. RESULTS: Forty-six patients were identified for analysis, all of whom were chemotherapy-naive. Only five patients treated in this cohort had NF1. BRAF fusion was identified in 65% (22/34) of tested tumors. Best therapy response was partial response in nine patients and stable disease in twenty-five patients. Twelve patients had progressive disease. One-year, 3-year, and 5-year EFS probabilities for all patients were 69.6%, 39.4%, and 34.5%, respectively. Nine patients had admissions for febrile neutropenia and seven patients experienced one delay in chemotherapy due to neutropenia. Only two patients had to discontinue this chemotherapy regimen because of treatment-related toxicities [carboplatin allergy (n = 1) and vinblastine neuropathy (n = 1)]. CONCLUSION: C/VBL achieves similar EFS rates to other single-agent and combination cytotoxic chemotherapy regimens for pediatric LGG with manageable toxicities.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Glioma/tratamento farmacológico , Adolescente , Neoplasias Encefálicas/patologia , Carboplatina/administração & dosagem , Criança , Pré-Escolar , Feminino , Seguimentos , Glioma/patologia , Humanos , Lactente , Recém-Nascido , Masculino , Gradação de Tumores , Estudos Retrospectivos , Taxa de Sobrevida , Vimblastina/administração & dosagem
4.
Pediatr Blood Cancer ; 67(9): e28426, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32614133

RESUMO

BACKGROUND: Relapse occurs in 50% of pediatric ependymoma cases and has poor prognosis. Few studies have investigated the clinical progress of relapsed disease, and treatment lacks a standardized approach. METHODS AND MATERIALS: We analyzed 302 pediatric ependymoma cases. Tumor, demographic, and treatment variables were investigated for association with relapse risk, time to recurrence, and survival after relapse. DNA methylation profiling was performed for 135/302 cases, and predominant subgroups were EPN_PFA (n = 95) and EPN_RELA (n = 24). Chromosome 1q status was ascertained for 185/302 cases by fluorescent in-situ hybridization (FISH), multiplex ligation-dependent probe amplification (MLPA), and DNA methylation profiles. RESULTS: Sixty-two percent of cases relapsed, with a median of two recurrences with no difference between posterior fossa and supratentorial locations (66% vs 55% relapse rate). One hundred seventeen (38%) cases relapsed within two years and five (2%) beyond 10 years. The late relapses were clinically heterogeneous. Tumor grade and treatment affected risk and time to relapse variably across subgroups. After relapse, surgery and irradiation delayed disease progression with a minimal impact on survival across the entire cohort. In the EPN_PFA and EPN_RELA groups, 1q gain was independently associated with relapse risk (subhazard ratio [SHR] 4.307, P = 0.027 and SHR 1.982, P = 0.010, respectively) while EPN_PFA had increased relapse risk compared with EPN_RELA (SHR = 0.394, P = 0.018). CONCLUSIONS: Recurrent pediatric ependymoma is an aggressive disease with poor outcomes, for which current treatments are inadequate. We report that chromosome 1q gain increases relapse risk in common molecular subgroups in children but a deeper understanding of the underlying biology at relapse and novel therapeutic approaches are urgently needed.


Assuntos
Neoplasias Encefálicas , Cromossomos Humanos Par 1 , Metilação de DNA , DNA de Neoplasias , Ependimoma , Recidiva Local de Neoplasia , Adolescente , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/terapia , Criança , Pré-Escolar , Cromossomos Humanos Par 1/genética , Cromossomos Humanos Par 1/metabolismo , DNA de Neoplasias/genética , DNA de Neoplasias/metabolismo , Ependimoma/genética , Ependimoma/metabolismo , Ependimoma/mortalidade , Ependimoma/terapia , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/metabolismo , Recidiva Local de Neoplasia/mortalidade , Recidiva Local de Neoplasia/terapia , Estudos Retrospectivos , Fatores de Risco
5.
Pediatr Blood Cancer ; 67(1): e28028, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31595628

RESUMO

BACKGROUND: The use of next-generation sequencing for fusion identification is being increasingly applied and aids our understanding of tumor biology. Some fusions are responsive to approved targeted agents, while others have future potential for therapeutic targeting. Although some pediatric central nervous system tumors may be cured with surgery alone, many require adjuvant therapy associated with acute and long-term toxicities. Identification of targetable fusions can shift the treatment paradigm toward earlier integration of molecularly targeted agents. METHODS: Patients diagnosed with glial, glioneuronal, and ependymal tumors between 2002 and 2019 were retrospectively reviewed for fusion testing. Testing was done primarily using the ArcherDx FusionPlex Solid Tumor panel, which assesses fusions in 53 genes. In contrast to many previously published series chronicling fusions in pediatric patients, we compared histological features and the tumor classification subtype with the specific fusion identified. RESULTS: We report 24 cases of glial, glioneuronal, or ependymal tumors from pediatric patients with identified fusions. With the exception of BRAF:KIAA1549 and pilocytic/pilomyxoid astrocytoma morphology, and possibly QKI-MYB and angiocentric glioma, there was not a strong correlation between histological features/tumor subtype and the specific fusion. We report the unusual fusions of PPP1CB-ALK, CIC-LEUTX, FGFR2-KIAA159, and MN1-CXXC5 and detail their morphological features. CONCLUSIONS: Fusion testing proved to be informative in a high percentage of cases. A large majority of fusion events in pediatric glial, glioneuronal, and ependymal tumors can be identified by relatively small gene panels.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Encefálicas/patologia , Ependimoma/patologia , Glioma/patologia , Neoplasias Neuroepiteliomatosas/patologia , Proteínas de Fusão Oncogênica/genética , Neoplasias Encefálicas/classificação , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/terapia , Criança , Pré-Escolar , Terapia Combinada , Ependimoma/classificação , Ependimoma/genética , Ependimoma/terapia , Feminino , Seguimentos , Glioma/classificação , Glioma/genética , Glioma/terapia , Humanos , Lactente , Masculino , Neoplasias Neuroepiteliomatosas/classificação , Neoplasias Neuroepiteliomatosas/genética , Neoplasias Neuroepiteliomatosas/terapia , Prognóstico , Estudos Retrospectivos
6.
Acta Neuropathol ; 135(5): 757-777, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29541918

RESUMO

Adamantinomatous craniopharyngiomas (ACPs) are clinically challenging tumours, the majority of which have activating mutations in CTNNB1. They are histologically complex, showing cystic and solid components, the latter comprised of different morphological cell types (e.g. ß-catenin-accumulating cluster cells and palisading epithelium), surrounded by a florid glial reaction with immune cells. Here, we have carried out RNA sequencing on 18 ACP samples and integrated these data with an existing ACP transcriptomic dataset. No studies so far have examined the patterns of gene expression within the different cellular compartments of the tumour. To achieve this goal, we have combined laser capture microdissection with computational analyses to reveal groups of genes that are associated with either epithelial tumour cells (clusters and palisading epithelium), glial tissue or immune infiltrate. We use these human ACP molecular signatures and RNA-Seq data from two ACP mouse models to reveal that cell clusters are molecularly analogous to the enamel knot, a critical signalling centre controlling normal tooth morphogenesis. Supporting this finding, we show that human cluster cells express high levels of several members of the FGF, TGFB and BMP families of secreted factors, which signal to neighbouring cells as evidenced by immunostaining against the phosphorylated proteins pERK1/2, pSMAD3 and pSMAD1/5/9 in both human and mouse ACP. We reveal that inhibiting the MAPK/ERK pathway with trametinib, a clinically approved MEK inhibitor, results in reduced proliferation and increased apoptosis in explant cultures of human and mouse ACP. Finally, we analyse a prominent molecular signature in the glial reactive tissue to characterise the inflammatory microenvironment and uncover the activation of inflammasomes in human ACP. We validate these results by immunostaining against immune cell markers, cytokine ELISA and proteome analysis in both solid tumour and cystic fluid from ACP patients. Our data support a new molecular paradigm for understanding ACP tumorigenesis as an aberrant mimic of natural tooth development and opens new therapeutic opportunities by revealing the activation of the MAPK/ERK and inflammasome pathways in human ACP.


Assuntos
Craniofaringioma/metabolismo , Sistema de Sinalização das MAP Quinases , Neoplasias Hipofisárias/metabolismo , Transcriptoma , Microambiente Tumoral/fisiologia , Animais , Biologia Computacional , Craniofaringioma/patologia , Craniofaringioma/terapia , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Inflamação/metabolismo , Inflamação/terapia , Microdissecção e Captura a Laser , Camundongos , Neuroglia/metabolismo , Odontogênese/fisiologia , Hipófise/embriologia , Hipófise/patologia , Neoplasias Hipofisárias/patologia , Neoplasias Hipofisárias/terapia , Análise de Sequência de RNA , Técnicas de Cultura de Tecidos
7.
Pediatr Blood Cancer ; 65(5): e26960, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29350470

RESUMO

BACKGROUND: A desperate need for novel therapies in pediatric ependymoma (EPN) exists, as chemotherapy remains ineffective and radiotherapy often fails. EPN have significant infiltration of immune cells, which correlates with outcome. Immune checkpoint inhibitors provide an avenue for new treatments. This study characterizes tumor-infiltrating immune cells in EPN and aims at predicting candidates for clinical trials using checkpoint inhibitors targeting PD-L1/PD-1 (programmed death ligand 1/programmed death 1). METHODS: The transcriptomic profiles of the primary study cohort of EPN and other pediatric brain tumors were interrogated to identify PD-L1 expression levels. Transcriptomic findings were validated using the western blotting, immunohistochemistry and flow cytometry. RESULTS: We evaluated PD-L1 mRNA expression across four intracranial subtypes of EPN in two independent cohorts and found supratentorial RELA fusion (ST-RELA) tumors to have significantly higher levels. There was a correlation between high gene expression and protein PD-L1 levels in ST-RELA tumors by both the western blot and immunohistochemisty. The investigation of EPN cell populations revealed PD-L1 was expressed on both tumor and myeloid cells in ST-RELA. Other subtypes had little PD-L1 in either tumor or myeloid cell compartments. Lastly, we measured PD-1 levels on tumor-infiltrating T cells and found ST-RELA tumors express PD-1 in both CD4 and CD8 T cells. A functional T-cell exhaustion assay found ST-RELA T cells to be exhausted and unable to secrete IFNγ on stimulation. CONCLUSIONS: These findings in ST-RELA suggest tumor evasion and immunsuppression due to PD-L1/PD-1-mediated T-cell exhaustion. Trials of checkpoint inhibitors in EPN should be enriched for ST-RELA tumors.


Assuntos
Antígeno B7-H1/metabolismo , Biomarcadores Tumorais/metabolismo , Ependimoma/metabolismo , Neoplasias Supratentoriais/metabolismo , Fator de Transcrição RelA/metabolismo , Adolescente , Adulto , Antígeno B7-H1/genética , Biomarcadores Tumorais/genética , Criança , Pré-Escolar , Estudos de Coortes , Ependimoma/genética , Ependimoma/patologia , Feminino , Seguimentos , Perfilação da Expressão Gênica , Humanos , Lactente , Masculino , Terapia de Alvo Molecular , Prognóstico , Neoplasias Supratentoriais/genética , Neoplasias Supratentoriais/patologia , Linfócitos T/metabolismo , Fator de Transcrição RelA/genética , Adulto Jovem
8.
Pediatr Blood Cancer ; 64(12)2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28548702

RESUMO

BACKGROUND: Posterior fossa (PF) ependymomas (EPNs) in infants less than 1 year of age (iEPN-PF) have a poorer clinical outcome than EPNs in older children. While radiation therapy is the standard of care for the latter, it is withheld in infants to avoid neurotoxicity to immature brain. It is unknown whether the adverse outcome in iEPN-PFs is due to treatment differences or aggressive biology. We examined this question using molecular profiling. METHODS: Six anaplastic iEPN-PFs were subjected to transcriptomic analysis and FISH for p16 loss and gains of 1q, and compared with anaplastic PF EPNs from older children. Results were validated by immunohistochemistry (IHC). RESULTS: All six iEPN-PFs were grouped within EPN PF subgroup A (PFA). E2F targets and G2M checkpoint were identified as the most enriched gene sets in iEPN-PF, which was validated in a larger independent cohort. Accordingly, MIB-1 IHC demonstrated a higher mitotic rate in iEPN-PFs than noninfant anaplastic EPN PFA. Genetic and protein analyses demonstrated that p16 loss and low p16 protein expression is a hallmark of iEPN-PF, and that none harbored 1q gains. Kaplan-Meier analysis confirmed the poorer clinical outcome of the iEPN-PF cohort. CONCLUSIONS: Biological differences, characterized by loss of p16 expression without gains of 1q in iEPN-PFs, as well as deregulated E2F target gene transcription, are indicative of deregulated p16-CDK4/6-pRB-E2F pathway activity. This may underlie the poor clinical outcome seen in this group of iEPN-PFs, rather than the withholding of radiation therapy. Results suggest a potential actionable therapy for iEPN-PF, namely cyclin-dependent kinase 4/6 (CDK4/6) inhibitors.


Assuntos
Neoplasias do Sistema Nervoso Central/genética , Fatores de Transcrição E2F/metabolismo , Ependimoma/genética , Genes p16 , Fatores Etários , Ciclo Celular , Neoplasias do Sistema Nervoso Central/terapia , Pré-Escolar , Ependimoma/terapia , Feminino , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Lactente , Estimativa de Kaplan-Meier , Masculino
9.
Pediatr Blood Cancer ; 64(11)2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28544128

RESUMO

BACKGROUND: Diffuse intrinsic pontine gliomas (DIPGs) and high-grade astrocytomas (HGA) continue to have dismal prognoses. The combination of cetuximab and irinotecan was demonstrated to be safe and tolerable in a previous pediatric phase 1 combination study. We developed this phase 2 trial to investigate the safety and efficacy of cetuximab given with radiation therapy followed by adjuvant cetuximab and irinotecan. METHODS: Eligible patients of age 3-21 years had newly diagnosed DIPG or HGA. Patients received radiation therapy (5,940 cGy) with concurrent cetuximab. Following radiation, patients received cetuximab weekly and irinotecan daily for 5 days per week for 2 weeks every 21 days for 30 weeks. Correlative studies were performed. The regimen was considered to be promising if the number of patients with 1-year progression-free survival (PFS) for DIPG and HGA was at least six of 25 and 14 of 26, respectively. RESULTS: Forty-five evaluable patients were enrolled (25 DIPG and 20 HGA). Six patients with DIPG and five with HGA were progression free at 1 year from the start of therapy with 1-year PFS of 29.6% and 18%, respectively. Fatigue, gastrointestinal complaints, electrolyte abnormalities, and rash were the most common adverse events and generally of grade 1 and 2. Increased epidermal growth factor receptor copy number but no K-ras mutations were identified in available samples. CONCLUSIONS: The trial did not meet the predetermined endpoint to deem this regimen successful for HGA. While the trial met the predetermined endpoint for DIPG, overall survival was not markedly improved from historical controls, therefore does not merit further study in this population.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Astrocitoma/terapia , Neoplasias do Tronco Encefálico/terapia , Quimiorradioterapia/mortalidade , Glioma/terapia , Adolescente , Adulto , Astrocitoma/patologia , Neoplasias do Tronco Encefálico/patologia , Camptotecina/administração & dosagem , Camptotecina/análogos & derivados , Cetuximab/administração & dosagem , Criança , Pré-Escolar , Terapia Combinada , Feminino , Seguimentos , Glioma/patologia , Humanos , Irinotecano , Masculino , Estadiamento de Neoplasias , Prognóstico , Taxa de Sobrevida , Adulto Jovem
10.
BMC Cancer ; 16: 647, 2016 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-27538997

RESUMO

BACKGROUND: Diffuse intrinsic pontine gliomas (DIPGs) are highly aggressive, fatal, childhood tumors that arise in the brainstem. DIPGs have no effective treatment, and their location and diffuse nature render them inoperable. Radiation therapy remains the only standard of care for this devastating disease. New therapeutic targets are needed to develop novel therapy for DIPG. METHODS: We examined the expression of PLK1 mRNA in DIPG tumor samples through microarray analysis and found it to be up regulated versus normal pons. Using the DIPG tumor cells, we inhibited PLK1 using a clinically relevant specific inhibitor BI 6727 and evaluated the effects on, proliferation, apoptosis, induction of DNA damage and radio sensitization of the DIPG tumor cells. RESULTS: Treatment of DIPG cell lines with BI 6727, a new generation, highly selective inhibitor of PLK1, resulted in decreased cell proliferation and a marked increase in cellular apoptosis. Cell cycle analysis showed a significant arrest in G2-M phase and a substantial increase in cell death. Treatment also resulted in an increased γH2AX expression, indicating induction of DNA damage. PLK1 inhibition resulted in radiosensitization of DIPG cells. CONCLUSION: These findings suggest that targeting PLK1 with small-molecule inhibitors, in combination with radiation therapy, will hold a novel strategy in the treatment of DIPG that warrants further investigation.


Assuntos
Neoplasias Encefálicas/genética , Proteínas de Ciclo Celular/genética , Glioma/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Proto-Oncogênicas/genética , Pteridinas/farmacologia , Radiossensibilizantes/farmacologia , Neoplasias Encefálicas/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Análise de Sequência de DNA/métodos , Regulação para Cima/efeitos dos fármacos , Quinase 1 Polo-Like
11.
J Immunol ; 191(9): 4880-8, 2013 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-24078694

RESUMO

Despite increasing evidence that antitumor immune control exists in the pediatric brain, these findings have yet to be exploited successfully in the clinic. A barrier to development of immunotherapeutic strategies in pediatric brain tumors is that the immunophenotype of these tumors' microenvironment has not been defined. To address this, the current study used multicolor FACS of disaggregated tumor to systematically characterize the frequency and phenotype of infiltrating immune cells in the most common pediatric brain tumor types. The initial study cohort consisted of 7 pilocytic astrocytoma (PA), 19 ependymoma (EPN), 5 glioblastoma (GBM), 6 medulloblastoma (MED), and 5 nontumor brain (NT) control samples obtained from epilepsy surgery. Immune cell types analyzed included both myeloid and T cell lineages and respective markers of activated or suppressed functional phenotypes. Immune parameters that distinguished each of the tumor types were identified. PA and EPN demonstrated significantly higher infiltrating myeloid and lymphoid cells compared with GBM, MED, or NT. Additionally, PA and EPN conveyed a comparatively activated/classically activated myeloid cell-skewed functional phenotype denoted in particular by HLA-DR and CD64 expression. In contrast, GBM and MED contained progressively fewer infiltrating leukocytes and more muted functional phenotypes similar to that of NT. These findings were recapitulated using whole tumor expression of corresponding immune marker genes in a large gene expression microarray cohort of pediatric brain tumors. The results of this cross-tumor comparative analysis demonstrate that different pediatric brain tumor types exhibit distinct immunophenotypes, implying that specific immunotherapeutic approaches may be most effective for each tumor type.


Assuntos
Neoplasias Encefálicas/classificação , Neoplasias Encefálicas/imunologia , Imunofenotipagem , Células Mieloides/imunologia , Linfócitos T/imunologia , Adolescente , Astrocitoma/imunologia , Encéfalo/imunologia , Neoplasias Encefálicas/genética , Criança , Estudos de Coortes , Ependimoma/imunologia , Epilepsia/imunologia , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Glioblastoma/imunologia , Antígenos HLA-DR/metabolismo , Humanos , Meduloblastoma/imunologia , Receptores de IgG/metabolismo , Microambiente Tumoral
12.
Mol Cancer ; 13: 72, 2014 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-24661910

RESUMO

BACKGROUND: Medulloblastoma is the most common type of malignant brain tumor that afflicts children. Although recent advances in chemotherapy and radiation have improved outcomes, high-risk patients do poorly with significant morbidity. METHODS: To identify new molecular targets, we performed an integrated genomic analysis using structural and functional methods. Gene expression profiling in 16 medulloblastoma patient samples and subsequent gene set enrichment analysis indicated that cell cycle-related kinases were associated with disease development. In addition a kinome-wide small interfering RNA (siRNA) screen was performed to identify kinases that, when inhibited, could prevent cell proliferation. The two genome-scale analyses were combined to identify key vulnerabilities in medulloblastoma. The inhibition of one of the identified targets was further investigated using RNAi and a small molecule inhibitor. RESULTS: Combining the two analyses revealed that mitosis-related kinases were critical determinants of medulloblastoma cell proliferation. RNA interference (RNAi)-mediated knockdown of WEE1 kinase and other mitotic kinases was sufficient to reduce medulloblastoma cell proliferation. These data prompted us to examine the effects of inhibiting WEE1 by RNAi and by a small molecule inhibitor of WEE1, MK-1775, in medulloblastoma cell lines. MK-1775 inhibited the growth of medulloblastoma cell lines, induced apoptosis and increased DNA damage at nanomolar concentrations. Further, MK-1775 was synergistic with cisplatin in reducing medulloblastoma cell proliferation and resulted in an associated increase in cell death. In vivo MK-1775 suppressed medulloblastoma tumor growth as a single agent. CONCLUSIONS: Taken together, these findings highlight mitotic kinases and, in particular, WEE1 as a rational therapeutic target for medulloblastoma.


Assuntos
Proteínas de Ciclo Celular/biossíntese , Meduloblastoma/genética , Terapia de Alvo Molecular , Proteínas Nucleares/biossíntese , Proteínas Tirosina Quinases/biossíntese , Apoptose/efeitos dos fármacos , Ciclo Celular/genética , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Pré-Escolar , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genoma Humano , Genômica , Humanos , Meduloblastoma/metabolismo , Meduloblastoma/patologia , Proteínas Nucleares/genética , Inibidores de Proteínas Quinases/administração & dosagem , Proteínas Tirosina Quinases/genética , Pirazóis/administração & dosagem , Pirimidinas/administração & dosagem , Pirimidinonas
13.
Acta Neuropathol ; 127(5): 731-45, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24240813

RESUMO

Better understanding of ependymoma (EPN) biology at relapse is needed to improve therapy at this critical event. Convincing data exist defining transcriptionally distinct posterior fossa (PF) sub-groups A and B at diagnosis. The clinical and biological consequence of these sub-groups at recurrence has not yet been defined. Genome and transcriptome microarray profiles and clinical variables of matched primary and first recurrent PF EPN pairs were used to identify biologically distinct patterns of progression between EPN sub-groups at recurrence. Key findings were validated by histology and immune function assays. Transcriptomic profiles were partially conserved at recurrence. However, 4 of 14 paired samples changed sub-groups at recurrence, and significant sub-group-specific transcriptomic changes between primary and recurrent tumors were identified, which were predominantly immune-related. Further examination revealed that Group A primary tumors harbor an immune gene signature and cellular functionality consistent with an immunosuppressive phenotype associated with tissue remodeling and wound healing. Conversely, Group B tumors develop an adaptive, antigen-specific immune response signature and increased T-cell infiltration at recurrence. Clinical distinctions between sub-groups become more apparent after first recurrence. Group A tumors were more often sub-totally resected and had a significantly shorter time to subsequent progression and worse overall survival. Minimal tumor-specific genomic changes were observed for either PF Groups A or B at recurrence. Molecular sub-groups of PF EPN convey distinct immunobiologic signatures at diagnosis and recurrence, providing potential biologic rationale to their disparate clinical outcomes. Immunotherapeutic approaches may be warranted, particularly in Group A PF EPN.


Assuntos
Ependimoma/diagnóstico , Ependimoma/imunologia , Neoplasias Infratentoriais/diagnóstico , Neoplasias Infratentoriais/imunologia , Recidiva Local de Neoplasia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Estudos de Coortes , Citocinas/metabolismo , Ependimoma/genética , Ependimoma/cirurgia , Feminino , Humanos , Imuno-Histoquímica , Neoplasias Infratentoriais/genética , Neoplasias Infratentoriais/cirurgia , Masculino , Análise em Microsséries , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Prognóstico , Transcriptoma , Adulto Jovem
14.
Pediatr Blood Cancer ; 61(1): 120-7, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23956023

RESUMO

BACKGROUND: Atypical teratoid rhabdoid tumors (AT-RT) are pediatric tumors of the central nervous system with limited treatment options and poor survival rate. We investigated whether enhancing chemotherapy toxicity by depleting intracellular glutathione (GSH; a key molecule in cisplatin resistance) with high dose acetaminophen (AAP), may improve therapeutic efficacy in AT-RT in vitro. PROCEDURE: BT16 (cisplatin-resistant) and BT12 (cisplatin-sensitive) AT-RT cell lines were treated with combinations of AAP, cisplatin, and the anti-oxidant N-acetylcysteine (NAC). Cell viability, GSH and peroxide concentrations, mitochondrial damage, and apoptosis were evaluated in vitro. RESULTS: AAP enhanced cisplatin cytotoxicity in cisplatin-resistant BT16 cells but not cisplatin-sensitive BT12 cells. Baseline GSH levels were elevated in BT16 cells compared to BT12 cells, and AAP decreased GSH to a greater magnitude in BT16 cells than BT12 cells. Unlike BT12 cells, BT16 cells did not have elevated peroxide levels upon treatment with cisplatin alone, but did have elevated levels when treated with AAP + cisplatin. Both cell lines had markedly increased mitochondrial injury when treated with AAP + cisplatin relative to either drug treatment alone. The enhanced toxic effects were partially reversed with concurrent administration of NAC. CONCLUSIONS: Our results suggest that AAP could be used as a chemo-enhancement agent to potentiate cisplatin chemotherapeutic efficacy particularly in cisplatin-resistant AT-RT tumors with high GSH levels in clinical settings.


Assuntos
Acetaminofen/administração & dosagem , Acetilcisteína/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Cisplatino/administração & dosagem , Tumor Rabdoide , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Glutationa/metabolismo , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Tumor Rabdoide/metabolismo , Tumor Rabdoide/patologia
15.
J Immunol ; 189(4): 1920-7, 2012 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-22802421

RESUMO

Survival in the majority of high-grade astrocytoma (HGA) patients is very poor, with only a rare population of long-term survivors. A better understanding of the biological factors associated with long-term survival in HGA would aid development of more effective therapy and survival prediction. Factors associated with long-term survival have not been extensively studied using unbiased genome-wide expression analyses. In the current study, gene expression microarray profiles of HGA from long-term survivors were interrogated for discovery of survival-associated biological factors. Ontology analyses revealed that increased expression of immune function-related genes was the predominant biological factor that positively correlated with longer survival. A notable T cell signature was present within this prognostic immune gene set. Using immune cell-specific gene classifiers, both T cell-associated and myeloid linage-associated genes were shown to be enriched in HGA from long-term versus short-term survivors. Association of immune function and cell-specific genes with survival was confirmed independently in a larger publicly available glioblastoma gene expression microarray data set. Histology was used to validate the results of microarray analyses in a larger cohort of long-term survivors of HGA. Multivariate analyses demonstrated that increased immune cell infiltration was a significant independent variable contributing to longer survival, as was Karnofsky/Lansky performance score. These data provide evidence of a prognostic anti-tumor adaptive immune response and rationale for future development of immunotherapy in HGA.


Assuntos
Astrocitoma/genética , Astrocitoma/imunologia , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/imunologia , Sobreviventes , Astrocitoma/mortalidade , Neoplasias Encefálicas/mortalidade , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Avaliação de Estado de Karnofsky , Linfócitos do Interstício Tumoral/imunologia , Análise de Sequência com Séries de Oligonucleotídeos , Prognóstico , Modelos de Riscos Proporcionais
16.
Neuro Oncol ; 26(3): 538-552, 2024 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-37934854

RESUMO

BACKGROUND: Pediatric high-grade gliomas (PHGG) are aggressive brain tumors with 5-year survival rates ranging from <2% to 20% depending upon subtype. PHGG presents differently from patient to patient and is intratumorally heterogeneous, posing challenges in designing therapies. We hypothesized that heterogeneity occurs because PHGG comprises multiple distinct tumor and immune cell types in varying proportions, each of which may influence tumor characteristics. METHODS: We obtained 19 PHGG samples from our institution's pediatric brain tumor bank. We constructed a comprehensive transcriptomic dataset at the single-cell level using single-cell RNA-Seq (scRNA-Seq), identified known glial and immune cell types, and performed differential gene expression and gene set enrichment analysis. We conducted multi-channel immunofluorescence (IF) staining to confirm the transcriptomic results. RESULTS: Our PHGG samples included 3 principal predicted tumor cell types: astrocytes, oligodendrocyte progenitors (OPCs), and mesenchymal-like cells (Mes). These cell types differed in their gene expression profiles, pathway enrichment, and mesenchymal character. We identified a macrophage population enriched in mesenchymal and inflammatory gene expression as a possible source of mesenchymal tumor characteristics. We found evidence of T-cell exhaustion and suppression. CONCLUSIONS: PHGG comprises multiple distinct proliferating tumor cell types. Microglia-derived macrophages may drive mesenchymal gene expression in PHGG. The predicted Mes tumor cell population likely derives from OPCs. The variable tumor cell populations rely on different oncogenic pathways and are thus likely to vary in their responses to therapy.


Assuntos
Neoplasias Encefálicas , Glioma , Humanos , Criança , Glioma/patologia , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Perfilação da Expressão Gênica , Sequenciamento do Exoma , Fenótipo
17.
bioRxiv ; 2024 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-38645202

RESUMO

Pediatric low-grade gliomas (pLGG) comprise 35% of all brain tumors. Despite favorable survival, patients experience significant morbidity from disease and treatments. A deeper understanding of pLGG biology is essential to identify novel, more effective, and less toxic therapies. We utilized single cell RNA sequencing (scRNA-seq), spatial transcriptomics, and cytokine analyses to characterize and understand tumor and immune cell heterogeneity across pLGG. scRNA-seq revealed tumor and immune cells within the tumor microenvironment (TME). Tumor cell subsets revealed a developmental hierarchy with progenitor and mature cell populations. Immune cells included myeloid and lymphocytic cells. There was a significant difference between the prevalence of two major myeloid subclusters between pilocytic astrocytoma (PA) and ganglioglioma (GG). Bulk and single-cell cytokine analyses evaluated the immune cell signaling cascade with distinct immune phenotypes among tumor samples. KIAA1549-BRAF tumors appeared more immunogenic, secreting higher levels of immune cell activators and chemokines, compared to BRAF V600E tumors. Spatial transcriptomics revealed the differential gene expression of these chemokines and their location within the TME. A multi-pronged analysis of pLGG demonstrated the complexity of the pLGG TME and differences between genetic drivers that may influence their response to immunotherapy. Further investigation of immune cell infiltration and tumor-immune interactions is warranted. Key points: There is a developmental hierarchy in neoplastic population comprising of both progenitor-like and mature cell types in both PA and GG.A more immunogenic, immune activating myeloid population is present in PA compared to GG. Functional analysis and spatial transcriptomics show higher levels of immune mobilizing chemokines in KIAA1549-BRAF fusion PA tumor samples compared to BRAF V600E GG samples. Importance of the Study: While scRNA seq provides information on cellular heterogeneity within the tumor microenvironment (TME), it does not provide a complete picture of how these cells are interacting or where they are located. To expand on this, we used a three-pronged approach to better understand the biology of pediatric low-grade glioma (pLGG). By analyzing scRNA-seq, secreted cytokines and spatial orientation of cells within the TME, we strove to gain a more complete picture of the complex interplay between tumor and immune cells within pLGG. Our data revealed a complex heterogeneity in tumor and immune populations and identified an interesting difference in the immune phenotype among different subtypes.

18.
Clin Cancer Res ; 30(8): 1544-1554, 2024 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-38334950

RESUMO

PURPOSE: There are no effective treatment strategies for children with highest-risk posterior fossa group A ependymoma (PFA). Chromosome 1q gains (1q+) are present in approximately 25% of newly diagnosed PFA tumors, and this number doubles at recurrence. Seventy percent of children with chromosome 1q+ PFA will die because of the tumor, highlighting the urgent need to develop new therapeutic strategies for this population. EXPERIMENTAL DESIGN: In this study, we utilize 1q+ PFA in vitro and in vivo models to test the efficacy of combination radiation and chemotherapy in a preclinical setting. RESULTS: 5-fluorouracil (5FU) enhances radiotherapy in 1q+ PFA cell lines. Specifically, 5FU increases p53 activity mediated by the extra copy of UCK2 located on chromosome 1q in 1q+ PFA. Experimental downregulation of UCK2 resulted in decreased 5FU sensitivity in 1q+ PFA cells. In in vitro studies, a combination of 5FU, retinoid tretinoin (ATRA), and radiation provided the greatest reduction in cellular proliferation and greatest increase in markers of apoptosis in 1q+ PFA cell lines compared with other treatment arms. Similarly, in vivo experiments demonstrated significant enhancement of survival in mice treated with combination radiation and 5FU and ATRA. CONCLUSIONS: These results are the first to identify a chromosome 1q+ specific therapy approach in 1q+ PFA. Existing phase I studies have already established single-agent pediatric safety and dosages of 5FU and ATRA, allowing for expedited clinical application as phase II trials for children with high-risk PFA.


Assuntos
Ependimoma , Neoplasias Infratentoriais , Criança , Humanos , Animais , Camundongos , Neoplasias Infratentoriais/genética , Neoplasias Infratentoriais/patologia , Neoplasias Infratentoriais/terapia , Resultado do Tratamento , Ependimoma/genética , Ependimoma/terapia , Fluoruracila , Cromossomos/metabolismo
19.
Neuro Oncol ; 26(6): 1109-1123, 2024 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-38334125

RESUMO

BACKGROUND: Cellular senescence can have positive and negative effects on the body, including aiding in damage repair and facilitating tumor growth. Adamantinomatous craniopharyngioma (ACP), the most common pediatric sellar/suprasellar brain tumor, poses significant treatment challenges. Recent studies suggest that senescent cells in ACP tumors may contribute to tumor growth and invasion by releasing a senesecence-associated secretory phenotype. However, a detailed analysis of these characteristics has yet to be completed. METHODS: We analyzed primary tissue samples from ACP patients using single-cell, single-nuclei, and spatial RNA sequencing. We performed various analyses, including gene expression clustering, inferred senescence cells from gene expression, and conducted cytokine signaling inference. We utilized LASSO to select essential gene expression pathways associated with senescence. Finally, we validated our findings through immunostaining. RESULTS: We observed significant diversity in gene expression and tissue structure. Key factors such as NFKB, RELA, and SP1 are essential in regulating gene expression, while senescence markers are present throughout the tissue. SPP1 is the most significant cytokine signaling network among ACP cells, while the Wnt signaling pathway predominantly occurs between epithelial and glial cells. Our research has identified links between senescence-associated features and pathways, such as PI3K/Akt/mTOR, MYC, FZD, and Hedgehog, with increased P53 expression associated with senescence in these cells. CONCLUSIONS: A complex interplay between cellular senescence, cytokine signaling, and gene expression pathways underlies ACP development. Further research is crucial to understand how these elements interact to create novel therapeutic approaches for patients with ACP.


Assuntos
Senescência Celular , Craniofaringioma , Aprendizado de Máquina , Neoplasias Hipofisárias , Humanos , Craniofaringioma/metabolismo , Craniofaringioma/patologia , Craniofaringioma/genética , Neoplasias Hipofisárias/patologia , Neoplasias Hipofisárias/metabolismo , Neoplasias Hipofisárias/genética , Biomarcadores Tumorais/metabolismo , Biomarcadores Tumorais/genética , Fenótipo , Regulação Neoplásica da Expressão Gênica , Criança , Masculino , Feminino
20.
Neurochem Res ; 38(9): 1838-49, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23743623

RESUMO

The antioxidant glutathione (GSH) plays a critical role in maintaining intracellular redox homeostasis but in tumors the GSH biosynthetic pathway is often dysregulated, contributing to tumor resistance to radiation and chemotherapy. Glutamate-cysteine ligase (GCL) catalyzes the first and rate-limiting reaction in GSH synthesis, and enzyme function is controlled by GSH feedback inhibition or by transcriptional upregulation of the catalytic (GCLC) and modifier (GCLM) subunits. However, it has recently been reported that the activity of GCLC and the formation of GCL can be modified by reactive aldehyde products derived from lipid peroxidation. Due to the susceptibility of GCLC to posttranslational modifications by reactive aldehydes, we examined the potential for 2-deoxy-D-ribose (2dDR) to glycate GCLC and regulate enzyme activity and GCL formation. 2dDR was found to directly modify both GCLC and GCLM in vitro, resulting in a significant inhibition of GCLC and GCL enzyme activity without altering substrate affinity or feedback inhibition. 2dDR-mediated glycation also inhibited GCL subunit heterodimerization and formation of the GCL holoenzyme complex while not causing dissociation of pre-formed holoenzyme. This PTM could be of particular importance in glioblastoma (GBM) where intratumoral necrosis provides an abundance of thymidine, which can be metabolized by thymidine phosphorylase (TP) to form 2dDR. TP is expressed at high levels in human GBM tumors and shRNA knockdown of TP in U87 GBM cells results in a significant increase in cellular GCL enzymatic activity.


Assuntos
Neoplasias Encefálicas/metabolismo , Desoxirribose/metabolismo , Resistencia a Medicamentos Antineoplásicos , Glioblastoma/metabolismo , Glucose/metabolismo , Glutamato-Cisteína Ligase/metabolismo , Neoplasias Encefálicas/enzimologia , Neoplasias Encefálicas/patologia , Catálise , Linhagem Celular Tumoral , Eletroforese em Gel de Poliacrilamida , Glioblastoma/enzimologia , Glioblastoma/patologia , Humanos , Cinética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
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