RESUMEN
Children with Group 3 medulloblastoma (G3 MB) have a very poor prognosis, and many do not survive beyond 5 years after diagnosis. A factor that may contribute to this is the lack of available targeted therapy. Expression of protein lin-28 homolog B (LIN28B), a regulator of developmental timing, is upregulated in several cancers, including G3 MB, and is associated with worse survival in this disease. Here, we investigate the role of the LIN28B pathway in G3 MB and demonstrate that the LIN28B-lethal-7 (let-7; a microRNA that is a tumor suppressor)-lymphokine-activated killer T-cell-originated protein kinase (PBK; also known as PDZ-binding kinase) axis promotes G3 MB proliferation. LIN28B knockdown in G3-MB-patient-derived cell lines leads to a significant reduction in cell viability and proliferation in vitro and in prolonged survival of mice with orthotopic tumors. The LIN28 inhibitor N-methyl-N-[3-(3-methyl-1,2,4-triazolo[4,3-b]pyridazin-6-yl)phenyl]acetamide (1632) significantly reduces G3 MB cell growth and demonstrates efficacy in reducing tumor growth in mouse xenograft models. Inhibiting PBK using HI-TOPK-032 also results in a significant reduction in G3 MB cell viability and proliferation. Together, these results highlight a critical role for the LIN28B-let-7-PBK pathway in G3 MB and provide preliminary preclinical results for drugs targeting this pathway.
Asunto(s)
Neoplasias Cerebelosas , Meduloblastoma , MicroARNs , Humanos , Ratones , Animales , Meduloblastoma/genética , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Proliferación Celular/genética , MicroARNs/genética , Neoplasias Cerebelosas/genética , Línea Celular Tumoral , Proteínas de Unión al ARN/genéticaRESUMEN
Ependymoma is a tumor of the brain or spinal cord. The two most common and aggressive molecular groups of ependymoma are the supratentorial ZFTA-fusion associated and the posterior fossa ependymoma group A. In both groups, tumors occur mainly in young children and frequently recur after treatment. Although molecular mechanisms underlying these diseases have recently been uncovered, they remain difficult to target and innovative therapeutic approaches are urgently needed. Here, we use genome-wide chromosome conformation capture (Hi-C), complemented with CTCF and H3K27ac ChIP-seq, as well as gene expression and DNA methylation analysis in primary and relapsed ependymoma tumors, to identify chromosomal conformations and regulatory mechanisms associated with aberrant gene expression. In particular, we observe the formation of new topologically associating domains ('neo-TADs') caused by structural variants, group-specific 3D chromatin loops, and the replacement of CTCF insulators by DNA hyper-methylation. Through inhibition experiments, we validate that genes implicated by these 3D genome conformations are essential for the survival of patient-derived ependymoma models in a group-specific manner. Thus, this study extends our ability to reveal tumor-dependency genes by 3D genome conformations even in tumors that lack targetable genetic alterations.
Asunto(s)
Ependimoma , Recurrencia Local de Neoplasia , Niño , Humanos , Preescolar , Recurrencia Local de Neoplasia/genética , Cromosomas , Mapeo Cromosómico , Ependimoma/genética , Ependimoma/patología , Genoma , Cromatina/genéticaRESUMEN
Major obstacles in brain cancer treatment include the blood-tumor barrier (BTB), which limits the access of most therapeutic agents, and quiescent tumor cells, which resist conventional chemotherapy. Here, we show that Sox2+ tumor cells project cellular processes to ensheathe capillaries in mouse medulloblastoma (MB), a process that depends on the mechanosensitive ion channel Piezo2. MB develops a tissue stiffness gradient as a function of distance to capillaries. Sox2+ tumor cells perceive substrate stiffness to sustain local intracellular calcium, actomyosin tension, and adhesion to promote cellular process growth and cell surface sequestration of ß-catenin. Piezo2 knockout reverses WNT/ß-catenin signaling states between Sox2+ tumor cells and endothelial cells, compromises the BTB, reduces the quiescence of Sox2+ tumor cells, and markedly enhances the MB response to chemotherapy. Our study reveals that mechanosensitive tumor cells construct the BTB to mask tumor chemosensitivity. Targeting Piezo2 addresses the BTB and tumor quiescence properties that underlie treatment failures in brain cancer.
Asunto(s)
Neoplasias Encefálicas , beta Catenina , Ratones , Animales , beta Catenina/metabolismo , beta Catenina/uso terapéutico , Células Endoteliales/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/patología , Encéfalo/metabolismo , Canales Iónicos/metabolismo , Barrera Hematoencefálica/metabolismoRESUMEN
Medulloblastoma (MB) comprises a group of heterogeneous paediatric embryonal neoplasms of the hindbrain with strong links to early development of the hindbrain1-4. Mutations that activate Sonic hedgehog signalling lead to Sonic hedgehog MB in the upper rhombic lip (RL) granule cell lineage5-8. By contrast, mutations that activate WNT signalling lead to WNT MB in the lower RL9,10. However, little is known about the more commonly occurring group 4 (G4) MB, which is thought to arise in the unipolar brush cell lineage3,4. Here we demonstrate that somatic mutations that cause G4 MB converge on the core binding factor alpha (CBFA) complex and mutually exclusive alterations that affect CBFA2T2, CBFA2T3, PRDM6, UTX and OTX2. CBFA2T2 is expressed early in the progenitor cells of the cerebellar RL subventricular zone in Homo sapiens, and G4 MB transcriptionally resembles these progenitors but are stalled in developmental time. Knockdown of OTX2 in model systems relieves this differentiation blockade, which allows MB cells to spontaneously proceed along normal developmental differentiation trajectories. The specific nature of the split human RL, which is destined to generate most of the neurons in the human brain, and its high level of susceptible EOMES+KI67+ unipolar brush cell progenitor cells probably predisposes our species to the development of G4 MB.
Asunto(s)
Diferenciación Celular , Neoplasias Cerebelosas , Meduloblastoma , Metencéfalo , Diferenciación Celular/genética , Linaje de la Célula , Neoplasias Cerebelosas/clasificación , Neoplasias Cerebelosas/genética , Neoplasias Cerebelosas/patología , Cerebelo/embriología , Cerebelo/patología , Subunidades alfa del Factor de Unión al Sitio Principal/genética , Proteínas Hedgehog/metabolismo , Histona Demetilasas , Humanos , Antígeno Ki-67/metabolismo , Meduloblastoma/clasificación , Meduloblastoma/genética , Meduloblastoma/patología , Metencéfalo/embriología , Metencéfalo/patología , Proteínas Musculares , Mutación , Factores de Transcripción Otx/deficiencia , Factores de Transcripción Otx/genética , Proteínas Represoras , Proteínas de Dominio T Box/metabolismo , Factores de TranscripciónRESUMEN
For decades, cell biologists and cancer researchers have taken advantage of non-murine species to increase our understanding of the molecular processes that drive normal cell and tissue development, and when perturbed, cause cancer. The advent of whole-genome sequencing has revealed the high genetic homology of these organisms to humans. Seminal studies in non-murine organisms such as Drosophila melanogaster, Caenorhabditis elegans, and Danio rerio identified many of the signaling pathways involved in cancer. Studies in these organisms offer distinct advantages over mammalian cell or murine systems. Compared to murine models, these three species have shorter lifespans, are less resource intense, and are amenable to high-throughput drug and RNA interference screening to test a myriad of promising drugs against novel targets. In this review, we introduce species-specific breeding strategies, highlight the advantages of modeling brain tumors in each non-mammalian species, and underscore the successes attributed to scientific investigation using these models. We conclude with an optimistic proposal that discoveries in the fields of cancer research, and in particular neuro-oncology, may be expedited using these powerful screening tools and strategies.
Asunto(s)
Neoplasias Encefálicas , Pez Cebra , Animales , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/genética , Caenorhabditis elegans , Drosophila melanogaster , Humanos , Ratones , Transducción de SeñalRESUMEN
Posterior fossa A (PFA) ependymomas are lethal malignancies of the hindbrain in infants and toddlers. Lacking highly recurrent somatic mutations, PFA ependymomas are proposed to be epigenetically driven tumors for which model systems are lacking. Here we demonstrate that PFA ependymomas are maintained under hypoxia, associated with restricted availability of specific metabolites to diminish histone methylation, and increase histone demethylation and acetylation at histone 3 lysine 27 (H3K27). PFA ependymomas initiate from a cell lineage in the first trimester of human development that resides in restricted oxygen. Unlike other ependymomas, transient exposure of PFA cells to ambient oxygen induces irreversible cellular toxicity. PFA tumors exhibit a low basal level of H3K27me3, and, paradoxically, inhibition of H3K27 methylation specifically disrupts PFA tumor growth. Targeting metabolism and/or the epigenome presents a unique opportunity for rational therapy for infants with PFA ependymoma.
Asunto(s)
Ependimoma/genética , Ependimoma/metabolismo , Epigenoma/genética , Neoplasias Infratentoriales/genética , Neoplasias Infratentoriales/metabolismo , Animales , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Línea Celular , Proliferación Celular/genética , Metilación de ADN/genética , Epigenómica/métodos , Histonas/genética , Histonas/metabolismo , Humanos , Lactante , Lisina/genética , Lisina/metabolismo , Masculino , Ratones Endogámicos C57BL , Mutación/genéticaRESUMEN
Recurrent medulloblastoma and ependymoma are universally lethal, with no approved targeted therapies and few candidates presently under clinical evaluation. Nearly all recurrent medulloblastomas and posterior fossa group A (PFA) ependymomas are located adjacent to and bathed by the cerebrospinal fluid, presenting an opportunity for locoregional therapy, bypassing the blood-brain barrier. We identify three cell-surface targets, EPHA2, HER2 and interleukin 13 receptor α2, expressed on medulloblastomas and ependymomas, but not expressed in the normal developing brain. We validate intrathecal delivery of EPHA2, HER2 and interleukin 13 receptor α2 chimeric antigen receptor T cells as an effective treatment for primary, metastatic and recurrent group 3 medulloblastoma and PFA ependymoma xenografts in mouse models. Finally, we demonstrate that administration of these chimeric antigen receptor T cells into the cerebrospinal fluid, alone or in combination with azacytidine, is a highly effective therapy for multiple metastatic mouse models of group 3 medulloblastoma and PFA ependymoma, thereby providing a rationale for clinical trials of these approaches in humans.
Asunto(s)
Neoplasias Encefálicas/terapia , Vacunas contra el Cáncer/administración & dosificación , Líquido Cefalorraquídeo/efectos de los fármacos , Ependimoma/terapia , Inmunoterapia Adoptiva/métodos , Meduloblastoma/terapia , Animales , Neoplasias Encefálicas/líquido cefalorraquídeo , Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/patología , Neoplasias Cerebelosas/líquido cefalorraquídeo , Neoplasias Cerebelosas/inmunología , Neoplasias Cerebelosas/patología , Neoplasias Cerebelosas/terapia , Líquido Cefalorraquídeo/inmunología , Niño , Preescolar , Sistemas de Liberación de Medicamentos/métodos , Ependimoma/líquido cefalorraquídeo , Ependimoma/inmunología , Ependimoma/patología , Femenino , Células HEK293 , Humanos , Lactante , Inyecciones Intraventriculares , Masculino , Meduloblastoma/líquido cefalorraquídeo , Meduloblastoma/inmunología , Meduloblastoma/patología , Ratones , Metástasis de la Neoplasia , Receptores Quiméricos de Antígenos/administración & dosificación , Receptores Quiméricos de Antígenos/genética , Receptores Quiméricos de Antígenos/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/trasplante , Resultado del Tratamiento , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Cancers are caused by genomic alterations known as drivers. Hundreds of drivers in coding genes are known but, to date, only a handful of noncoding drivers have been discovered-despite intensive searching1,2. Attention has recently shifted to the role of altered RNA splicing in cancer; driver mutations that lead to transcriptome-wide aberrant splicing have been identified in multiple types of cancer, although these mutations have only been found in protein-coding splicing factors such as splicing factor 3b subunit 1 (SF3B1)3-6. By contrast, cancer-related alterations in the noncoding component of the spliceosome-a series of small nuclear RNAs (snRNAs)-have barely been studied, owing to the combined challenges of characterizing noncoding cancer drivers and the repetitive nature of snRNA genes1,7,8. Here we report a highly recurrent A>C somatic mutation at the third base of U1 snRNA in several types of tumour. The primary function of U1 snRNA is to recognize the 5' splice site via base-pairing. This mutation changes the preferential A-U base-pairing between U1 snRNA and the 5' splice site to C-G base-pairing, and thus creates novel splice junctions and alters the splicing pattern of multiple genes-including known drivers of cancer. Clinically, the A>C mutation is associated with heavy alcohol use in patients with hepatocellular carcinoma, and with the aggressive subtype of chronic lymphocytic leukaemia with unmutated immunoglobulin heavy-chain variable regions. The mutation in U1 snRNA also independently confers an adverse prognosis to patients with chronic lymphocytic leukaemia. Our study demonstrates a noncoding driver in spliceosomal RNAs, reveals a mechanism of aberrant splicing in cancer and may represent a new target for treatment. Our findings also suggest that driver discovery should be extended to a wider range of genomic regions.
Asunto(s)
Mutación , Neoplasias/genética , ARN Nuclear Pequeño/genética , Empalmosomas/genética , Humanos , Neoplasias/patología , Neoplasias/fisiopatología , Sitios de Empalme de ARN , Empalme del ARN , Factores de Empalme de ARN/genéticaRESUMEN
In cancer, recurrent somatic single-nucleotide variants-which are rare in most paediatric cancers-are confined largely to protein-coding genes1-3. Here we report highly recurrent hotspot mutations (r.3A>G) of U1 spliceosomal small nuclear RNAs (snRNAs) in about 50% of Sonic hedgehog (SHH) medulloblastomas. These mutations were not present across other subgroups of medulloblastoma, and we identified these hotspot mutations in U1 snRNA in only <0.1% of 2,442 cancers, across 36 other tumour types. The mutations occur in 97% of adults (subtype SHHδ) and 25% of adolescents (subtype SHHα) with SHH medulloblastoma, but are largely absent from SHH medulloblastoma in infants. The U1 snRNA mutations occur in the 5' splice-site binding region, and snRNA-mutant tumours have significantly disrupted RNA splicing and an excess of 5' cryptic splicing events. Alternative splicing mediated by mutant U1 snRNA inactivates tumour-suppressor genes (PTCH1) and activates oncogenes (GLI2 and CCND2), and represents a target for therapy. These U1 snRNA mutations provide an example of highly recurrent and tissue-specific mutations of a non-protein-coding gene in cancer.
Asunto(s)
Neoplasias Cerebelosas/genética , Proteínas Hedgehog/genética , Meduloblastoma/genética , ARN Nuclear Pequeño/genética , Adolescente , Adulto , Empalme Alternativo , Proteínas Hedgehog/metabolismo , Humanos , Mutación , Sitios de Empalme de ARN , Empalme del ARNRESUMEN
BACKGROUND: Inflammatory bowel diseases are associated with increased expression of zinc-dependent Matrix Metalloproteinase 9 (MMP-9). A stark dysregulation of intestinal mucosal homeostasis has been observed in patients with chronic inflammatory bowel diseases. We therefore sought to determine the contribution of MMP-9 to the pathogenesis of Citrobacter rodentium-induced colitis and its effects on gut microbiome homeostasis. RESULTS: Wild-type and MMP-9-/- mice aged 5-6 weeks were challenged with C. rodentium by orogastric gavage and sacrificed either 10 or 30 days post-infection. Disease severity was assessed by histological analysis of colonic epithelial hyperplasia and by using an in vivo intestinal permeability assay. Changes in the inflammatory responses were measured by using qPCR, and the composition of the fecal microbiome evaluated with both qPCR and terminal restriction fragment length polymorphism. Activation and localization of MMP-9 to the apical surface of the colonic epithelium in response to C. rodentium infection was demonstrated by both zymography and immunocytochemistry. The pro-inflammatory response to infection, including colonic epithelial cell hyperplasia and barrier dysfunction, was similar, irrespective of genotype. Nonmetric multidimensional scaling of terminal restriction fragments revealed a different fecal microbiome composition and C. rodentium colonization pattern between genotypes, with MMP-9-/- having elevated levels of protective segmented filamentous bacteria and interleukin-17, and lower levels of C. rodentium. MMP-9-/- but not wild-type mice were also protected from reductions in fecal microbial diversity in response to the bacterial enteric infection. CONCLUSIONS: These results demonstrate that MMP-9 expression in the colon causes alterations in the fecal microbiome and has an impact on the pathogenesis of bacterial-induced colitis in mice.