RESUMEN
Diffuse intrinsic pontine glioma (DIPG) is an incurable brain tumor of childhood characterized by histone mutations at lysine 27, which results in epigenomic dysregulation. There has been a failure to develop effective treatment for this tumor. Using a combined RNAi and chemical screen targeting epigenomic regulators, we identify the polycomb repressive complex 1 (PRC1) component BMI1 as a critical factor for DIPG tumor maintenance in vivo. BMI1 chromatin occupancy is enriched at genes associated with differentiation and tumor suppressors in DIPG cells. Inhibition of BMI1 decreases cell self-renewal and attenuates tumor growth due to induction of senescence. Prolonged BMI1 inhibition induces a senescence-associated secretory phenotype, which promotes tumor recurrence. Clearance of senescent cells using BH3 protein mimetics co-operates with BMI1 inhibition to enhance tumor cell killing in vivo.
Asunto(s)
Envejecimiento/genética , Glioma Pontino Intrínseco Difuso/genética , Complejo Represivo Polycomb 1/metabolismo , Astrocitoma/genética , Neoplasias del Tronco Encefálico/tratamiento farmacológico , Neoplasias del Tronco Encefálico/genética , Diferenciación Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Niño , Preescolar , Cromatina/genética , Glioma Pontino Intrínseco Difuso/tratamiento farmacológico , Glioma Pontino Intrínseco Difuso/metabolismo , Epigenómica , Femenino , Glioma/tratamiento farmacológico , Glioma/genética , Glioma/patología , Histonas/metabolismo , Humanos , Lisina/metabolismo , Masculino , Mutación , Recurrencia Local de Neoplasia/tratamiento farmacológico , Recurrencia Local de Neoplasia/genética , Complejo Represivo Polycomb 1/antagonistas & inhibidores , Complejo Represivo Polycomb 1/genéticaRESUMEN
Pineoblastomas (PBs) are rare, aggressive pediatric brain tumors of the pineal gland with modest overall survival despite intensive therapy. We sought to define the clinical and molecular spectra of PB to inform new treatment approaches for this orphan cancer. Tumor, blood, and clinical data from 91 patients with PB or supratentorial primitive neuroectodermal tumor (sPNETs/CNS-PNETs), and 2 pineal parenchymal tumors of intermediate differentiation (PPTIDs) were collected from 29 centres in the Rare Brain Tumor Consortium. We used global DNA methylation profiling to define a core group of PB from 72/93 cases, which were delineated into five molecular sub-groups. Copy number, whole exome and targeted sequencing, and miRNA expression analyses were used to evaluate the clinico-pathologic significance of each sub-group. Tumors designated as group 1 and 2 almost exclusively exhibited deleterious homozygous loss-of-function alterations in miRNA biogenesis genes (DICER1, DROSHA, and DGCR8) in 62 and 100% of group 1 and 2 tumors, respectively. Recurrent alterations of the oncogenic MYC-miR-17/92-RB1 pathway were observed in the RB and MYC sub-group, respectively, characterized by RB1 loss with gain of miR-17/92, and recurrent gain or amplification of MYC. PB sub-groups exhibited distinct clinical features: group 1-3 arose in older children (median ages 5.2-14.0 years) and had intermediate to excellent survival (5-year OS of 68.0-100%), while Group RB and MYC PB patients were much younger (median age 1.3-1.4 years) with dismal survival (5-year OS 37.5% and 28.6%, respectively). We identified age < 3 years at diagnosis, metastatic disease, omission of upfront radiation, and chr 16q loss as significant negative prognostic factors across all PBs. Our findings demonstrate that PB exhibits substantial molecular heterogeneity with sub-group-associated clinical phenotypes and survival. In addition to revealing novel biology and therapeutics, molecular sub-grouping of PB can be exploited to reduce treatment intensity for patients with favorable biology tumors.
Asunto(s)
Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Glándula Pineal , Pinealoma/genética , Pinealoma/patología , Adolescente , Adulto , Factores de Edad , Neoplasias Encefálicas/mortalidad , Niño , Preescolar , Estudios de Cohortes , Femenino , Humanos , Lactante , Masculino , MicroARNs/metabolismo , Mutación/genética , Pinealoma/mortalidad , Sistema de Registros , Tasa de Supervivencia , Adulto JovenRESUMEN
Loss of SMARCB1 is the hallmark genetic event that characterizes rhabdoid tumors in children. Rhabdoid tumors of the brain (ATRT) occur in young children and are particularly challenging with poor long-term survival. SMARCB1 is a member of the SWI/SNF chromatin remodeling complex that is responsible for determining cellular pluripotency and lineage commitment. The mechanisms by which SMARCB1 deletion results in tumorigenesis remain unclear. Recent studies demonstrate that ATRT consists of 3 genomic subgroups with a subset of poor outcome tumors expressing high BMP and MYC pathway activation. Here we show that MYC occupies distinct promoter loci in ATRT compared to embryonic stem (ES) cells. Furthermore, using human ATRT cell lines, patient-derived cell culture, ex vivo patient-derived tumor, and orthotopic xenograft models, we show that MYC inhibition is a molecular vulnerability in SMARCB1-deleted tumors and that such inhibition effectively suppresses BMP and pluripotency-associated genomic programs, attenuates tumor cell self-renewal, promotes senescence, and inhibits ATRT tumor growth in vivo. Transgenic expression of Omomyc (a bona-fide MYC dominant negative) or chemical inhibition of MYC transcriptomic programs with the BET inhibitor JQ1 phenocopy genetic depletion of MYC, effectively restricting ATRT tumor growth and opening a promising therapeutic avenue for rhabdoid tumors in children.
Asunto(s)
Transformación Celular Neoplásica/genética , Regulación Neoplásica de la Expresión Génica/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , Tumor Rabdoide/genética , Proteína SMARCB1/genética , Teratoma/genética , Animales , Azepinas/farmacología , Proteínas Morfogenéticas Óseas/genética , Proteínas Morfogenéticas Óseas/metabolismo , Línea Celular Tumoral , Autorrenovación de las Células/efectos de los fármacos , Autorrenovación de las Células/genética , Senescencia Celular/efectos de los fármacos , Senescencia Celular/genética , Cromatina/genética , Cromatina/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Ratones , Ratones Desnudos , Proteínas Proto-Oncogénicas c-myc/genética , Tumor Rabdoide/patología , Teratoma/patología , Triazoles/farmacología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Hypoxia and expression of hypoxia-related biomarkers are associated with disease progression and treatment failure in prostate cancer (PCa). We have reported that exosomes (nanovesicles of 30-150 nm in diameter) secreted by human PCa cells under hypoxia promote invasiveness and stemness in naïve PCa cells. Here, we identified the unique microRNAs (miRNAs) loaded in exosomes secreted by PCa cells under hypoxia. Using TaqMan® array microRNA cards, we analyzed the miRNA profile in exosomes secreted by human PCa LNCaP cells under hypoxic (ExoHypoxic) and normoxic (ExoNormoxic) conditions. We identified 292 miRNAs loaded in both ExoHypoxic and ExoNormoxic. The top 11 miRNAs with significantly higher level in ExoHypoxic compared to ExoNormoxic were miR-517a, miR-204, miR-885, miR-143, miR-335, miR-127, miR-542, miR-433, miR-451, miR-92a and miR-181a; and top nine miRNA with significantly lower expression level in ExoHypoxic compared to ExoNormoxic were miR-521, miR-27a, miR-324, miR-579, miR-502, miR-222, miR-135b, miR-146a and miR-491. Importantly, the two differentially expressed miRNAs miR-885 (increased expression) and miR-521 (decreased expression) showed similar expression pattern in exosomes isolated from the serum of PCa patients compared to healthy individuals. Additionally, miR-204 and miR-222 displayed correlated expression patterns in prostate tumors (Pearson R = 0.66, p < 0.0001) by The Cancer Genome Atlas (TCGA) prostate adenocarcinoma (PRAD) genomic dataset analysis. Overall, the present study identified unique miRNAs with differential expression in exosomes secreted from hypoxic PCa cells and suggests their potential usefulness as a biomarker of hypoxia in PCa patients.
RESUMEN
Atypical teratoid rhabdoid tumor (ATRT) is an aggressive and malignant pediatric brain tumor. Polo-like kinase 1 (PLK1) is highly expressed in many cancers and essential for mitosis. Overexpression of PLK1 promotes chromosome instability and aneuploidy by overriding the G2-M DNA damage and spindle checkpoints. Recent studies suggest that targeting PLK1 by small molecule inhibitors is a promising approach to tumor therapy. We investigated the effect of PLK1 inhibition in ATRT. Gene expression analysis showed that PLK1 was overexpressed in ATRT patient samples and tumor cell lines. Genetic inhibition of PLK1 with shRNA potently suppressed ATRT cell growth in vitro. Treatment with the PLK1 inhibitor BI 6727 (Volasertib) significantly decreased cell growth, inhibited clonogenic potential, and induced apoptosis. BI6727 treatment led to G2-M phase arrest, consistent with PLK1's role as a critical regulator of mitosis. Moreover, inhibition of PLK1 by BI6727 suppressed the tumor-sphere formation of ATRT cells. Treatment also significantly decreased levels of the DNA damage proteins Ku80 and RAD51 and increased γ-H2AX expression, indicating that BI 6727 can induce DNA damage. Importantly, BI6727 significantly enhanced radiation sensitivity of ATRT cells. In vivo, BI6727 slowed growth of ATRT tumors and prolonged survival in a xenograft model. PLK1 inhibition is a compelling new therapeutic approach for treating ATRT, and the use of BI6727 should be evaluated in clinical studies.
RESUMEN
Kinase inhibitors are effective cancer therapies, but tumors frequently develop resistance. Current strategies to circumvent resistance target the same or parallel pathways. We report here that targeting a completely different process, autophagy, can overcome multiple BRAF inhibitor resistance mechanisms in brain tumors. BRAFV600Emutations occur in many pediatric brain tumors. We previously reported that these tumors are autophagy-dependent and a patient was successfully treated with the autophagy inhibitor chloroquine after failure of the BRAFV600E inhibitor vemurafenib, suggesting autophagy inhibition overcame the kinase inhibitor resistance. We tested this hypothesis in vemurafenib-resistant brain tumors. Genetic and pharmacological autophagy inhibition overcame molecularly distinct resistance mechanisms, inhibited tumor cell growth, and increased cell death. Patients with resistance had favorable clinical responses when chloroquine was added to vemurafenib. This provides a fundamentally different strategy to circumvent multiple mechanisms of kinase inhibitor resistance that could be rapidly tested in clinical trials in patients with BRAFV600E brain tumors.
Asunto(s)
Antineoplásicos/uso terapéutico , Autofagia/efectos de los fármacos , Neoplasias Encefálicas/tratamiento farmacológico , Cloroquina/uso terapéutico , Resistencia a Antineoplásicos , Indoles/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/metabolismo , Sulfonamidas/uso terapéutico , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Humanos , Proteínas Proto-Oncogénicas B-raf/genética , Resultado del Tratamiento , VemurafenibRESUMEN
We recently reported that atypical teratoid rhabdoid tumors (ATRTs) comprise at least two transcriptional subtypes with different clinical outcomes; however, the mechanisms underlying therapeutic heterogeneity remained unclear. In this study, we analyzed 191 primary ATRTs and 10 ATRT cell lines to define the genomic and epigenomic landscape of ATRTs and identify subgroup-specific therapeutic targets. We found ATRTs segregated into three epigenetic subgroups with distinct genomic profiles, SMARCB1 genotypes, and chromatin landscape that correlated with differential cellular responses to a panel of signaling and epigenetic inhibitors. Significantly, we discovered that differential methylation of a PDGFRB-associated enhancer confers specific sensitivity of group 2 ATRT cells to dasatinib and nilotinib, and suggest that these are promising therapies for this highly lethal ATRT subtype.
Asunto(s)
Neoplasias del Sistema Nervioso Central/genética , Cromatina/genética , Epigenómica/métodos , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/genética , Tumor Rabdoide/genética , Proteína SMARCB1/genética , Teratoma/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Neoplasias del Sistema Nervioso Central/tratamiento farmacológico , Metilación de ADN , Dasatinib/farmacología , Dasatinib/uso terapéutico , Epigénesis Genética/efectos de los fármacos , Humanos , Mutación , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/antagonistas & inhibidores , Tumor Rabdoide/tratamiento farmacológico , Teratoma/tratamiento farmacológicoRESUMEN
Medulloblastoma is the most common type of malignant brain tumor that affects children. Although recent advances in chemotherapy and radiation have improved outcomes, high-risk patients perform poorly with significant morbidity. Gene expression profiling has revealed that monopolar spindle 1 (MPS1) (TTK1) is highly expressed in medulloblastoma patient samples compared to that noted in normal cerebellum. MPS1 is a key regulator of the spindle assembly checkpoint (SAC), a mitotic mechanism specifically required for proper chromosomal alignment and segregation. The SAC can be activated in aneuploid cancer cells and MPS1 is overexpressed in many types of cancers. A previous study has demonstrated the effectiveness of inhibiting MPS1 with small-molecule inhibitors, but the role of MPS1 in medulloblastoma is unknown. In the present study, we demonstrated that MPS1 inhibition by shRNA or with a small-molecule drug, NMS-P715, resulted in decreased cell growth, inhibition of clonogenic potential and induction of apoptosis in cells belonging to both the Shh and group 3 medulloblastoma genomic signature. These findings highlight MPS1 as a rational therapeutic target for medulloblastoma.
Asunto(s)
Apoptosis/efectos de los fármacos , Proteínas de Ciclo Celular/biosíntesis , Meduloblastoma/genética , Terapia Molecular Dirigida , Proteínas Serina-Treonina Quinasas/biosíntesis , Proteínas Tirosina Quinasas/biosíntesis , Ciclo Celular/efectos de los fármacos , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Ciclo Celular/genética , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HeLa , Humanos , Puntos de Control de la Fase M del Ciclo Celular/efectos de los fármacos , Meduloblastoma/tratamiento farmacológico , Meduloblastoma/patología , Mitosis/genética , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/genética , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/genética , Pirazoles/administración & dosificación , Quinazolinas/administración & dosificaciónRESUMEN
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.
Asunto(s)
Neoplasias Encefálicas/genética , Proteínas de Ciclo Celular/genética , Glioma/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/genética , Pteridinas/farmacología , Fármacos Sensibilizantes a Radiaciones/farmacología , Neoplasias Encefálicas/tratamiento farmacológico , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Daño del ADN , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Análisis de Secuencia de ADN/métodos , Regulación hacia Arriba/efectos de los fármacos , Quinasa Tipo Polo 1RESUMEN
Checkpoint kinase 1 (CHK1) is an integral component of the cell cycle as well as the DNA Damage Response (DDR) pathway. Previous work has demonstrated the effectiveness of inhibiting CHK1 with small-molecule inhibitors, but the role of CHK1 mediated DDR in medulloblastoma is unknown. CHK1, both at the mRNA and protein level, is highly expressed in medulloblastoma and elevated CHK1 expression in Group3 medulloblastoma is an adverse prognostic marker. CHK1 inhibition with the small-molecule drug AZD7762, results in decreased cell growth, increased DNA damage and cell apoptosis. Furthermore, AZD7762 acts in synergy with cisplatin in reducing cell proliferation in medulloblastoma. Similar phenotypic changes were observed with another CHK1 inhibitor, PF477736, as well as genetic knockdown using siRNA against CHK1. Treatments with small-molecule inhibitors of CHK1 profoundly modulated the expression of both upstream and downstream target proteins within the CHK1 signaling pathways. This suggests the presence of a feedback loop in activating CHK1. Overall, our results demonstrate that small-molecule inhibition of CHK1 in combination with, cisplatin, is more advantageous than either treatment alone, especially for Group 3 medulloblastoma, and therefore this combined therapeutic approach serves as an avenue for further investigation.
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Biomarcadores de Tumor/análisis , Neoplasias Cerebelosas/patología , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1)/biosíntesis , Meduloblastoma/patología , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Apoptosis/efectos de los fármacos , Benzodiazepinonas/farmacología , Biomarcadores de Tumor/metabolismo , Supervivencia Celular/efectos de los fármacos , Neoplasias Cerebelosas/enzimología , Neoplasias Cerebelosas/mortalidad , Cisplatino/farmacología , Supervivencia sin Enfermedad , Genes myc , Humanos , Estimación de Kaplan-Meier , Meduloblastoma/enzimología , Meduloblastoma/mortalidad , Pronóstico , Pirazoles/farmacología , Tiofenos/farmacología , Urea/análogos & derivados , Urea/farmacologíaRESUMEN
Ependymoma (EPN) in childhood is a brain tumor with substantial mortality. Inflammatory response has been identified as a molecular signature of high-risk Group A EPN. To better understand the biology of this phenotype and aid therapeutic development, transcriptomic data from Group A and B EPN patient tumor samples, and additional malignant and normal brain data, were analyzed to identify the mechanism underlying EPN Group A inflammation. Enrichment of IL6 and STAT3 pathway genes were found to distinguish Group A EPN from Group B EPN and other brain tumors, implicating an IL6 activation of STAT3 mechanism. EPN tumor cell growth was shown to be dependent on STAT3 activity, as demonstrated using shRNA knockdown and pharmacologic inhibition of STAT3 that blocked proliferation and induced apoptosis. The inflammatory factors secreted by EPN tumor cells were shown to reprogram myeloid cells, and this paracrine effect was characterized by a significant increase in pSTAT3 and IL8 secretion. Myeloid polarization was shown to be dependent on tumor secretion of IL6, and these effects could be reversed using IL6-neutralizing antibody or IL6 receptor-targeted therapeutic antibody tocilizumab. Polarized myeloid cell production of IL8 drove unpolarized myeloid cells to upregulate CD163 and to produce a number of proinflammatory cytokines. Collectively, these findings indicate that constitutive IL6/STAT3 pathway activation is important in driving tumor growth and inflammatory cross-talk with myeloid cells within the Group A EPN microenvironment. Effective design of Group A-targeted therapy for children with EPN may require reversal of this potentially immunosuppressive and protumor pathway.
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Ependimoma/metabolismo , Ependimoma/patología , Interleucina-6/metabolismo , Fenotipo , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Citocinas/biosíntesis , Ependimoma/genética , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Mediadores de Inflamación/metabolismo , Receptores de Lipopolisacáridos/metabolismo , Monocitos/metabolismo , Células Mieloides/metabolismo , FosforilaciónRESUMEN
INTRODUCTION: Pediatric adamantinomatous craniopharyngioma (ACP) is a histologically benign but clinically aggressive brain tumor that arises from the sellar/suprasellar region. Despite a high survival rate with current surgical and radiation therapy (75-95 % at 10 years), ACP is associated with debilitating visual, endocrine, neurocognitive and psychological morbidity, resulting in excheptionally poor quality of life for survivors. Identification of an effective pharmacological therapy could drastically decrease morbidity and improve long term outcomes for children with ACP. RESULTS: Using mRNA microarray gene expression analysis of 15 ACP patient samples, we have found several pharmaceutical targets that are significantly and consistently overexpressed in our panel of ACP relative to other pediatric brain tumors, pituitary tumors, normal pituitary and normal brain tissue. Among the most highly expressed are several targets of the kinase inhibitor dasatinib - LCK, EPHA2 and SRC; EGFR pathway targets - AREG, EGFR and ERBB3; and other potentially actionable cancer targets - SHH, MMP9 and MMP12. We confirm by western blot that a subset of these targets is highly expressed in ACP primary tumor samples. CONCLUSIONS: We report here the first published transcriptome for ACP and the identification of targets for rational therapy. Experimental drugs targeting each of these gene products are currently being tested clinically and pre-clinically for the treatment of other tumor types. This study provides a rationale for further pre-clinical and clinical studies of novel pharmacological treatments for ACP. Development of mouse and cell culture models for ACP will further enable the translation of these targets from the lab to the clinic, potentially ushering in a new era in the treatment of ACP.
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Craneofaringioma/metabolismo , Sistemas de Liberación de Medicamentos/métodos , Familia de Proteínas EGF/metabolismo , Receptores ErbB/metabolismo , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/metabolismo , Calidad de Vida/psicología , ARN Mensajero/metabolismo , Receptor EphA2/metabolismo , Receptor ErbB-3/metabolismo , Familia-src Quinasas/metabolismo , Adolescente , Anfirregulina , Niño , Preescolar , Craneofaringioma/tratamiento farmacológico , Craneofaringioma/genética , Familia de Proteínas EGF/genética , Receptores ErbB/genética , Femenino , Expresión Génica , Humanos , Lactante , Recién Nacido , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/genética , Masculino , Análisis por Micromatrices/métodos , Receptor EphA2/genética , Receptor ErbB-3/genética , Regulación hacia Arriba , Familia-src Quinasas/genéticaRESUMEN
BACKGROUND: Rhabdoid brain tumours, also called atypical teratoid rhabdoid tumours, are lethal childhood cancers with characteristic genetic alterations of SMARCB1/hSNF5. Lack of biological understanding of the substantial clinical heterogeneity of these tumours restricts therapeutic advances. We integrated genomic and clinicopathological analyses of a cohort of patients with atypical teratoid rhabdoid tumours to find out the molecular basis for clinical heterogeneity in these tumours. METHODS: We obtained 259 rhabdoid tumours from 37 international institutions and assessed transcriptional profiles in 43 primary tumours and copy number profiles in 38 primary tumours to discover molecular subgroups of atypical teratoid rhabdoid tumours. We used gene and pathway enrichment analyses to discover group-specific molecular markers and did immunohistochemical analyses on 125 primary tumours to evaluate clinicopathological significance of molecular subgroup and ASCL1-NOTCH signalling. FINDINGS: Transcriptional analyses identified two atypical teratoid rhabdoid tumour subgroups with differential enrichment of genetic pathways, and distinct clinicopathological and survival features. Expression of ASCL1, a regulator of NOTCH signalling, correlated with supratentorial location (p=0·004) and superior 5-year overall survival (35%, 95% CI 13-57, and 20%, 6-34, for ASCL1-positive and ASCL1-negative tumours, respectively; p=0·033) in 70 patients who received multimodal treatment. ASCL1 expression also correlated with superior 5-year overall survival (34%, 7-61, and 9%, 0-21, for ASCL1-positive and ASCL1-negative tumours, respectively; p=0·001) in 39 patients who received only chemotherapy without radiation. Cox hazard ratios for overall survival in patients with differential ASCL1 enrichment treated with chemotherapy with or without radiation were 2·02 (95% CI 1·04-3·85; p=0·038) and 3·98 (1·71-9·26; p=0·001). Integrated analyses of molecular subgroupings with clinical prognostic factors showed three distinct clinical risk groups of tumours with different therapeutic outcomes. INTERPRETATION: An integration of clinical risk factors and tumour molecular groups can be used to identify patients who are likely to have improved long-term radiation-free survival and might help therapeutic stratification of patients with atypical teratoid rhabdoid tumours. FUNDING: C17 Research Network, Genome Canada, b.r.a.i.n.child, Mitchell Duckman, Tal Doron and Suri Boon foundations.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/biosíntesis , Genómica , Receptores Notch/biosíntesis , Tumor Rabdoide/genética , Teratoma/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Niño , Preescolar , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Inmunohistoquímica , Lactante , Masculino , Pronóstico , Receptores Notch/genética , Tumor Rabdoide/patología , Factores de Riesgo , Transducción de Señal/genética , Teratoma/patologíaRESUMEN
Medulloblastoma is a pediatric brain tumor with a variable prognosis due to clinical and genomic heterogeneity. Among the 4 major genomic sub-groups, patients with MYC amplified tumors have a particularly poor prognosis despite therapy with surgery, radiation and chemotherapy. Targeting the MYC oncogene has traditionally been problematic. Here we report that MYC driven medulloblastoma can be targeted by inhibition of the bromodomain protein BRD4. We show that bromodomain inhibition with JQ1 restricts c-MYC driven transcriptional programs in medulloblastoma, suppresses medulloblastoma cell growth and induces a cell cycle arrest. Importantly JQ1 suppresses stem cell associated signaling in medulloblastoma cells and inhibits medulloblastoma tumor cell self-renewal. Additionally JQ1 also promotes senescence in medulloblastoma cells by activating cell cycle kinase inhibitors and inhibiting activity of E2F1. Furthermore BRD4 inhibition displayed an anti-proliferative, pro-senescence effect in a medulloblastoma model in vivo. In clinical samples we found that transcriptional programs suppressed by JQ1 are associated with adverse risk in medulloblastoma patients. Our work indicates that BRD4 inhibition attenuates stem cell signaling in MYC driven medulloblastoma and demonstrates the feasibility BET domain inhibition as a therapeutic approach in vivo.
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Proliferación Celular , Neoplasias Cerebelosas/prevención & control , Meduloblastoma/prevención & control , Células Madre Neoplásicas/patología , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-myc/metabolismo , Factores de Transcripción/antagonistas & inhibidores , Animales , Apoptosis , Azepinas/farmacología , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Western Blotting , Ciclo Celular , Proteínas de Ciclo Celular , Neoplasias Cerebelosas/genética , Neoplasias Cerebelosas/mortalidad , Neoplasias Cerebelosas/patología , Niño , Perfilación de la Expresión Génica , Humanos , Meduloblastoma/genética , Meduloblastoma/mortalidad , Meduloblastoma/patología , Ratones , Ratones Desnudos , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Pronóstico , Proteínas Proto-Oncogénicas c-myc/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-myc/genética , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tasa de Supervivencia , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Triazoles/farmacología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
UNLABELLED: Autophagy inhibition is a potential therapeutic strategy in cancer, but it is unknown which tumors will benefit. The BRAF(V600E) mutation has been identified as important in pediatric central nervous system (CNS) tumors and is known to affect autophagy in other tumor types. We evaluated CNS tumor cells with BRAF(V600E) and found that mutant (but not wild-type) cells display high rates of induced autophagy, are sensitive to pharmacologic and genetic autophagy inhibition, and display synergy when the clinically used autophagy inhibitor chloroquine was combined with the RAF inhibitor vemurafenib or standard chemotherapeutics. Importantly, we also demonstrate that chloroquine can improve vemurafenib sensitivity in a resistant ex vivo primary culture and provide the first demonstration in a patient harboring the V600E mutation treated with vemurafenib that the addition of chloroquine can improve clinical outcomes. These findings suggest that CNS tumors with BRAF(V600E) are autophagy-dependent and should be targeted with autophagy inhibition in combination with other therapeutic strategies. SIGNIFICANCE: Autophagy inhibition may improve cancer therapy, but it is unclear which tumors will benefit. We found that BRAF mutations cause brain tumor cells to depend on autophagy and display selective chemosensitization with autophagy inhibition. We present a pediatric case in which deliberate autophagy inhibition halted tumor growth and overcame acquired BRAF-inhibition resistance.
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Antineoplásicos/farmacología , Autofagia/efectos de los fármacos , Neoplasias Encefálicas/tratamiento farmacológico , Sistema Nervioso Central/efectos de los fármacos , Cloroquina/farmacología , Indoles/farmacología , Proteínas Proto-Oncogénicas B-raf/genética , Sulfonamidas/farmacología , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Sistema Nervioso Central/patología , Niño , Resistencia a Antineoplásicos/efectos de los fármacos , Sinergismo Farmacológico , Humanos , Mutación , VemurafenibRESUMEN
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.
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Proteínas de Ciclo Celular/biosíntesis , Meduloblastoma/genética , Terapia Molecular Dirigida , Proteínas Nucleares/biosíntesis , Proteínas Tirosina Quinasas/biosíntesis , Apoptosis/efectos de los fármacos , Ciclo Celular/genética , Proteínas de Ciclo Celular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Preescolar , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genoma Humano , Genómica , Humanos , Meduloblastoma/metabolismo , Meduloblastoma/patología , Proteínas Nucleares/genética , Inhibidores de Proteínas Quinasas/administración & dosificación , Proteínas Tirosina Quinasas/genética , Pirazoles/administración & dosificación , Pirimidinas/administración & dosificación , PirimidinonasRESUMEN
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.
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Ependimoma/diagnóstico , Ependimoma/inmunología , Neoplasias Infratentoriales/diagnóstico , Neoplasias Infratentoriales/inmunología , Recurrencia Local de Neoplasia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Estudios de Cohortes , Citocinas/metabolismo , Ependimoma/genética , Ependimoma/cirugía , Femenino , Humanos , Inmunohistoquímica , Neoplasias Infratentoriales/genética , Neoplasias Infratentoriales/cirugía , Masculino , Análisis por Micromatrices , Persona de Mediana Edad , Polimorfismo de Nucleótido Simple , Pronóstico , Transcriptoma , Adulto JovenRESUMEN
Brainstem gangliogliomas (GGs), often cannot be resected, have a much poorer prognosis than those located in more common supratentorial sites and may benefit from novel therapeutic approaches. Therapeutically targetable BRAF c.1799T>A (p.V600E) (BRAF(V600E) ) mutations are harbored in roughly 50% of collective GGs taken from all anatomical sites. Large numbers of pediatric brainstem GGs, however, have not been specifically assessed and anatomic-and age-restricted assessment of genetic and biological factors are becoming increasingly important. Pediatric brainstem GGs (n = 13), non-brainstem GGs (n = 11) and brainstem pilocytic astrocytomas (PAs) (n = 8) were screened by standard Sanger DNA sequencing of BRAF exon 15. Five of 13 (38%) pediatric GG harbored a definitive BRAF(V600E) mutation, with two others exhibiting an equivocal result by this method. BRAF(V600E) was also seen in five of 11 (45%) non-brainstem GGs and one of eight (13%) brainstem PAs. VE1 immunostaining for BRAF(V600E) showed concordance with sequencing in nine of nine brainstem GGs including the two cases equivocal by Sanger. The equivocal brainstem GGs were subsequently shown to harbor BRAF(V600E) using a novel, more sensitive, RNA-sequencing approach, yielding a final BRAF(V600E) mutation frequency of 54% (seven of 13) in brainstem GGs. BRAF(V600E) -targeted therapeutics should be a consideration for the high percentage of pediatric brainstem GGs refractory to conventional therapies.
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Neoplasias del Tronco Encefálico/genética , Ganglioglioma/genética , Proteínas Proto-Oncogénicas B-raf/genética , Adolescente , Neoplasias del Tronco Encefálico/patología , Niño , Preescolar , Exones , Femenino , Ganglioglioma/patología , Humanos , Lactante , Recién Nacido , Masculino , Mutación , Estudios Retrospectivos , Adulto JovenRESUMEN
BACKGROUND: Although, substantial experimental evidence related to diagnosis and treatment of pediatric central nervous system (CNS) neoplasms have been demonstrated, the understanding of the etiology and pathogenesis of the disease remains scarce. Recent microRNA (miRNA)-based research reveals the involvement of miRNAs in various aspects of CNS development and proposes that they might compose key molecules underlying oncogenesis. The current study evaluated miRNA differential expression detected between pediatric embryonal brain tumors and normal controls to characterize candidate biomarkers related to diagnosis, prognosis and therapy. METHODS: Overall, 19 embryonal brain tumors; 15 Medulloblastomas (MBs) and 4 Atypical Teratoid/Rabdoid Tumors (AT/RTs) were studied. As controls, 13 samples were used; The First-Choice Human Brain Reference RNA and 12 samples from deceased children who underwent autopsy and were not present with any brain malignancy. RNA extraction was carried out using the Trizol method, whilst miRNA extraction was performed with the mirVANA miRNA isolation kit. The experimental approach included miRNA microarrays covering 1211 miRNAs. Quantitative Real-Time Polymerase Chain Reaction was performed to validate the expression profiles of miR-34a and miR-601 in all 32 samples initially screened with miRNA microarrays and in an additional independent cohort of 30 patients (21MBs and 9 AT/RTs). Moreover, meta-analyses was performed in total 27 embryonal tumor samples; 19 MBs, 8 ATRTs and 121 control samples. Twelve germinomas were also used as an independent validation cohort. All deregulated miRNAs were correlated to patients' clinical characteristics and pathological measures. RESULTS: In several cases, there was a positive correlation between individual miRNA expression levels and laboratory or clinical characteristics. Based on that, miR-601 could serve as a putative tumor suppressor gene, whilst miR-34a as an oncogene. In general, miR-34a demonstrated oncogenic roles in all pediatric embryonal CNS neoplasms studied. CONCLUSIONS: Deeper understanding of the aberrant miRNA expression in pediatric embryonal brain tumors might aid in the development of tumor-specific miRNA signatures, which could potentially afford promising biomarkers related to diagnosis, prognosis and patient targeted therapy.
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Neoplasias del Sistema Nervioso Central/genética , MicroARNs/biosíntesis , Neoplasias de Células Germinales y Embrionarias/genética , Adolescente , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias del Sistema Nervioso Central/metabolismo , Neoplasias Cerebelosas/genética , Neoplasias Cerebelosas/metabolismo , Niño , Preescolar , Progresión de la Enfermedad , Femenino , Humanos , Masculino , Meduloblastoma/genética , Meduloblastoma/metabolismo , MicroARNs/genética , Neoplasias de Células Germinales y Embrionarias/metabolismo , Valor Predictivo de las Pruebas , Pronóstico , Teratoma/genética , Teratoma/metabolismo , Resultado del TratamientoRESUMEN
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.