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Strategies to overcome irreversible cochlear hair cell (HC) damage and loss in mammals are of vital importance to hearing recovery in patients with permanent hearing loss. In mature mammalian cochlea, co-activation of Myc and Notch1 reprograms supporting cells (SC) and promotes HC regeneration. Understanding of the underlying mechanisms may aid the development of a clinically relevant approach to achieve HC regeneration in the nontransgenic mature cochlea. By single-cell RNAseq, we show that MYC/NICD "rejuvenates" the adult mouse cochlea by activating multiple pathways including Wnt and cyclase activator of cyclic AMP (cAMP), whose blockade suppresses HC-like cell regeneration despite Myc/Notch activation. We screened and identified a combination (the cocktail) of drug-like molecules composing of small molecules and small interfering RNAs to activate the pathways of Myc, Notch1, Wnt and cAMP. We show that the cocktail effectively replaces Myc and Notch1 transgenes and reprograms fully mature wild-type (WT) SCs for HC-like cells regeneration in vitro. Finally, we demonstrate the cocktail is capable of reprogramming adult cochlea for HC-like cells regeneration in WT mice with HC loss in vivo. Our study identifies a strategy by a clinically relevant approach to reprogram mature inner ear for HC-like cells regeneration, laying the foundation for hearing restoration by HC regeneration.
Assuntos
Orelha Interna , Células Ciliadas Auditivas , Camundongos , Animais , Proliferação de Células/fisiologia , Células Ciliadas Auditivas/fisiologia , Orelha Interna/metabolismo , Cóclea/fisiologia , Regeneração/fisiologia , MamíferosRESUMO
SWI/SNF chromatin remodeling enzymes are multisubunit complexes that contain one of two catalytic subunits, BRG1 or BRM and 9-11 additional subunits called BRG1 or BRM-associated factors (BAFs). BRG1 interacts with the microphthalmia-associated transcription factor (MITF) and is required for melanocyte development in vitro and in vivo. The subunits of SWI/SNF that mediate interactions between BRG1 and MITF have not been elucidated. Three mutually exclusive isoforms of a 60-kDa subunit (BAF60A, B, or C) often facilitate interactions with transcription factors during lineage specification. We tested the hypothesis that a BAF60 subunit promotes interactions between MITF and the BRG1-containing SWI/SNF complex. We found that MITF can physically interact with BAF60A, BAF60B, and BAF60C. The interaction between MITF and BAF60A required the basic helix-loop-helix domain of MITF. Recombinant BAF60A pulled down recombinant MITF, suggesting that the interaction can occur in the absence of other SWI/SNF subunits and other transcriptional regulators of the melanocyte lineage. Depletion of BAF60A in differentiating melanoblasts inhibited melanin synthesis and expression of MITF target genes. MITF promoted BAF60A recruitment to melanocyte-specific promoters, and BAF60A was required to promote BRG1 recruitment and chromatin remodeling. Thus, BAF60A promotes interactions between MITF and the SWI/SNF complex and is required for melanocyte differentiation.
Assuntos
Diferenciação Celular , Proteínas Cromossômicas não Histona/metabolismo , DNA Helicases/metabolismo , Melanócitos/citologia , Melanócitos/metabolismo , Fator de Transcrição Associado à Microftalmia/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Ciclo Celular , Diferenciação Celular/genética , Regulação da Expressão Gênica , Células HEK293 , Humanos , Melaninas/biossíntese , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Fator de Transcrição Associado à Microftalmia/química , Modelos Biológicos , Oxirredutases/genética , Oxirredutases/metabolismo , Regiões Promotoras Genéticas/genética , Ligação Proteica , Subunidades Proteicas/metabolismoRESUMO
Brahma (BRM) and Brahma-related gene 1(BRG1) are catalytic subunits of SWItch/sucrose non-fermentable (SWI/SNF) chromatin remodeling complexes. BRM is epigenetically silenced in a wide-range of tumors. Mutations in the v-raf murine sarcoma viral oncogene homolog B1 (BRAF) gene occur frequently in melanoma and lead to constitutive activation of the mitogen-activated protein kinase (MAPK)/extracellular signal regulated kinase (ERK1/2) pathway. We tested the hypothesis that BRM expression is modulated by oncogenic BRAF and phosphorylation of ERK1/2 in melanocytes and melanoma cells. Expression of oncogenic BRAF in melanocytes and melanoma cells that are wild-type for BRAF decreased BRM expression and increased BRG1 expression. Inhibition of mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) or selective inhibition of BRAF in melanoma cells that harbor oncogenic BRAF increased BRM expression and decreased BRG1 expression. Increased BRM expression was associated with increased histone acetylation on the BRM promoter. Over-expression of BRM in melanoma cells that harbor oncogenic BRAF promoted changes in cell cycle progression and apoptosis consistent with a tumor suppressive role. Upon inhibition of BRAF(V600E) with PLX4032, BRM promoted survival. PLX4032 induced changes in BRM function were correlated with increased acetylation of the BRM protein. This study provides insights into the epigenetic consequences of inhibiting oncogenic BRAF in melanoma through modulation of SWI/SNF subunit expression and function.
Assuntos
Sistema de Sinalização das MAP Quinases , Melanoma/genética , Melanoma/metabolismo , Fatores de Transcrição/genética , Substituição de Aminoácidos , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , DNA Helicases/genética , Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Histonas/metabolismo , Humanos , Melanócitos/citologia , Melanócitos/metabolismo , Melanoma/patologia , Mutação , Proteínas Nucleares/genética , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas B-raf/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , RNA Neoplásico/metabolismo , RNA Interferente Pequeno/genética , Proteínas Recombinantes/genética , Proteína do Retinoblastoma/metabolismo , Fatores de Transcrição/antagonistas & inibidoresRESUMO
The transposase-accessible chromatin using sequencing (ATAC-seq) offers a simplified approach to detect chromatin changes in cancer cells after genetic intervention and drug treatment. Here, we present an optimized ATAC-seq protocol to elucidate chromatin accessibility changes at the epigenetic level in head and neck squamous cell carcinoma cells. We describe steps for cell lysate preparation, transposition, and tagmentation, followed by library amplification and purification. We then detail next-generation sequencing and data analysis. For complete details on the use and execution of this protocol, please refer to Buenrostro et al.,1 Chen et al.,2.
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Long non-coding RNAs (lncRNAs) are transcripts without coding potential that are pervasively expressed from the genome and have been increasingly reported to play crucial roles in all aspects of cell biology. They have been also heavily implicated in cancer development and progression, with both oncogenic and tumor suppressor functions. In this work, we identified and characterized a novel lncRNA, TAZ-AS202, expressed from the TAZ genomic locus and exerting pro-oncogenic functions in non-small cell lung cancer. TAZ-AS202 expression is under the control of YAP/TAZ-containing transcriptional complexes. We demonstrated that TAZ-AS202 is overexpressed in lung cancer tissue, compared with surrounding lung epithelium. In lung cancer cell lines TAZ-AS202 promotes cell migration and cell invasion. TAZ-AS202 regulates the expression of a set of genes belonging to cancer-associated pathways, including WNT and EPH-Ephrin signaling. The molecular mechanism underlying TAZ-AS202 function does not involve change of TAZ expression or activity, but increases the protein level of the transcription factor E2F1, which in turn regulates the expression of a large set of target genes, including the EPHB2 receptor. Notably, the silencing of both E2F1 and EPHB2 recapitulates TAZ-AS202 silencing cellular phenotype, indicating that they are essential mediators of its activity. Overall, this work unveiled a new regulatory mechanism that, by increasing E2F1 protein, modifies the non-small cell lung cancer cells transcriptional program, leading to enhanced aggressiveness features. The TAZ-AS202/E2F1/EPHB2 axis may be the target for new therapeutic strategies.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , RNA Longo não Codificante , Humanos , Neoplasias Pulmonares/patologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Carcinoma Pulmonar de Células não Pequenas/genética , Fator de Transcrição E2F1/genética , Fator de Transcrição E2F1/metabolismo , Efrinas/genética , Efrinas/metabolismo , Linhagem Celular Tumoral , Pulmão/metabolismo , Regulação Neoplásica da Expressão Gênica/genéticaRESUMO
This study investigates the role of integrin ß4 (ITGB4) and stemness-associated factor SOX2 in platinum resistance in lung squamous cell carcinoma (LUSC). The expression of SOX2 and ITGB4 is found to be high in all LUSC subtypes, but the impact of ITGB4 expression on overall patient survival varies by subtype. Cancer stem cells (CSCs) isolated from LUSC patients were found to be resistant to cisplatin, but knocking down ITGB4 or SOX2 sensitized them to cisplatin. Carfilzomib (CFZ) synergized with cisplatin and suppressed CSC growth by inhibiting ITGB4 and SOX2 expression. Additionally, CFZ was found to inhibit SOX2 expression epigenetically by inhibiting histone acetylation at the SOX2 promoter site. CFZ also suppressed the growth of SOX2-dependent small cell lung cancer cells in vitro and in vivo. The study highlights the unique function of CFZ as a transcriptional suppressor of SOX2, independent of its proteasome inhibitory function.
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The human airway contains specialized rare epithelial cells whose roles in respiratory disease are not well understood. Ionocytes express the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR), while chemosensory tuft cells express asthma-associated alarmins. However, surprisingly, exceedingly few mature tuft cells have been identified in human lung cell atlases despite the ready identification of rare ionocytes and neuroendocrine cells. To identify human rare cell progenitors and define their lineage relationship to mature tuft cells, we generated a deep lung cell atlas containing 311,748 single cell RNA-Seq (scRNA-seq) profiles from discrete anatomic sites along the large and small airways and lung lobes of explanted donor lungs that could not be used for organ transplantation. Of 154,222 airway epithelial cells, we identified 687 ionocytes (0.45%) that are present in similar proportions in both large and small airways, suggesting that they may contribute to both large and small airways pathologies in CF. In stark contrast, we recovered only 3 mature tuft cells (0.002%). Instead, we identified rare bipotent progenitor cells that can give rise to both ionocytes and tuft cells, which we termed tuft-ionocyte progenitor cells (TIP cells). Remarkably, the cycling fraction of these TIP cells was comparable to that of basal stem cells. We used scRNA-seq and scATAC-seq to predict transcription factors that mark this novel rare cell progenitor population and define intermediate states during TIP cell lineage transitions en route to the differentiation of mature ionocytes and tuft cells. The default lineage of TIP cell descendants is skewed towards ionocytes, explaining the paucity of mature tuft cells in the human airway. However, Type 2 and Type 17 cytokines, associated with asthma and CF, diverted the lineage of TIP cell descendants in vitro , resulting in the differentiation of mature tuft cells at the expense of ionocytes. Consistent with this model of mature tuft cell differentiation, we identify mature tuft cells in a patient who died from an asthma flare. Overall, our findings suggest that the immune signaling pathways active in asthma and CF may skew the composition of disease-relevant rare cells and illustrate how deep atlases are required for identifying physiologically-relevant scarce cell populations.
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With the introduction of immunotherapy, significant improvement has been made in the treatment of head and neck squamous cell carcinoma (HNSCC). However, only a small subset of patients with HNSCC benefit from immunotherapy. The current biomarker, a programmed cell death protein ligand 1 (PD-L1) expression that is widely used in treatment decision making for advanced HNSCC, has only a moderate predictive value. Additionally, PD-L1-based assay has critical inherent limitations due to its highly dynamic nature and lack of standardization. With the advance in molecular techniques and our understanding of biology, more reliable, reproducible, and practical novel biomarkers are being developed. These include but are not limited to neoantigen/mutation characteristics, immune transcriptomes, tumor-infiltrating immune cell composition, cancer epigenomic, proteomics and metabolic characteristics, and plasma-based and organoid assays.
Assuntos
Neoplasias de Cabeça e Pescoço , Inibidores de Checkpoint Imunológico , Antígeno B7-H1 , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Imunoterapia/métodos , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológicoRESUMO
Head and neck squamous cell carcinoma (HNSCC) and its treatments are associated with substantial morbidity, often resulting in cosmetic deformity and loss of physiologic functions including speech and swallowing. Despite advancements in treatment, 5-year survival rates for mucosal malignancies remain below 70%. Effective prevention of HNSCC demands an understanding of the molecular pathways of carcinogenesis. Specifically, defining features of pre-cancerous dysplastic lesions that indicate a better or worse prognosis is necessary to help identify patients who are likely to develop a carcinoma and allow a more aggressive approach to management. There remains a need for identification of biomarkers that can provide both early prognostic and predictive value in clinical decision-making by serving as both therapeutic targets as well as predictors of therapy response. Here, we comprehensively review the most frequently altered molecular biomarkers of malignant transformation in head and neck dysplasia. These markers are involved in a wide range of cellular processes in head and neck carcinogenesis, including extracellular matrix degradation, cell motility and invasion, cell-cell adhesion, solute transport, immortalization, metabolism, the cell cycle and apoptosis, transcription, and cell signaling.
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Analysis of The Cancer Genome Atlas and other published data of head and neck squamous cell carcinoma (HNSCC) reveals somatic alterations of the Hippo-YAP pathway in approximately 50% of HNSCC. Better strategies to target the YAP1 transcriptional complex are sought. Here, we show that FAT1, an upstream inhibitor of YAP1, is mutated either by missense or by truncating mutation in 29% of HNSCC. Comprehensive proteomic and drug-screening studies across pan-cancer models confirm that FAT1-mutant HNSCC exhibits selective and higher sensitivity to BRD4 inhibition by JQ1. Epigenomic analysis reveals an active chromatin state in FAT1-mutant HNSCC cells that is driven by the YAP/TAZ transcriptional complex through recruitment of BRD4 to deposit active histone marks, thereby maintaining an oncogenic transcriptional state. This study reveals a detailed cooperative mechanism between YAP1 and BRD4 in HNSCC and suggests a specific therapeutic opportunity for the treatment of this subset of head and neck cancer patients.
Assuntos
Proteínas de Ciclo Celular , Neoplasias de Cabeça e Pescoço , Proteínas Nucleares , Fatores de Transcrição , Proteínas de Sinalização YAP , Carcinogênese/genética , Carcinogênese/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Cromatina , Neoplasias de Cabeça e Pescoço/genética , Humanos , Proteínas Nucleares/genética , Proteômica , Carcinoma de Células Escamosas de Cabeça e Pescoço , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas de Sinalização YAP/genética , Proteínas de Sinalização YAP/metabolismoRESUMO
Advances in the treatment of malignant pleural mesothelioma (MPM) have been disappointing, despite the apparent need for new therapeutic options for this rare and devastating cancer. Drug resistance is common and surgical intervention has brought benefits only to a subset of patients. MPM is a heterogenous disease with a surprisingly low mutation rate and recent sequencing efforts have confirmed alterations in a limited number of tumor suppressors that do not provide apparent insights into the molecular mechanisms that drive this malignancy. There is increasing evidence that epigenetic regulation leads to immune evasion and transformation in MPM. Further, the low efficacy of immune checkpoint inhibitors is consistent with a suppression of genes involved in the anti-tumor immune response. We review three promising emerging therapeutic targets (STAT3, KDM4A, heparanase) and highlight their potential effects on the immune response.
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PURPOSE: While chemotherapy remains the standard treatment for triple-negative breast cancer (TNBC), identifying and managing chemoresistant tumors has proven elusive. We sought to discover hallmarks and therapeutically actionable features of refractory TNBC through molecular analysis of primary chemoresistant TNBC specimens. EXPERIMENTAL DESIGN: We performed transcriptional profiling of tumors from a phase II clinical trial of platinum chemotherapy for advanced TNBC (TBCRC-009), revealing a gene expression signature that identified de novo chemorefractory tumors. We then employed pharmacogenomic data mining, proteomic and other molecular studies to define the therapeutic vulnerabilities of these tumors. RESULTS: We reveal the RAS-GTPase-activating protein (RAS-GAP) RASAL2 as an upregulated factor that mediates chemotherapy resistance but also an exquisite collateral sensitivity to combination MAP kinase kinase (MEK1/2) and EGFR inhibitors in TNBC. Mechanistically, RASAL2 GAP activity is required to confer kinase inhibitor sensitivity, as RASAL2-high TNBCs sustain basal RAS activity through suppression of negative feedback regulators SPRY1/2, together with EGFR upregulation. Consequently, RASAL2 expression results in failed feedback compensation upon co-inhibition of MEK1/2 and EGFR that induces synergistic apoptosis in vitro and in vivo. In patients with TNBC, high RASAL2 levels predict clinical chemotherapy response and long-term outcomes, and are associated via direct transcriptional regulation with activated oncogenic Yes-Associated Protein (YAP). Accordingly, chemorefractory patient-derived TNBC models exhibit YAP activation, high RASAL2 expression, and tumor regression in response to MEK/EGFR inhibitor combinations despite well-tolerated intermittent dosing. CONCLUSIONS: These findings identify RASAL2 as a mediator of TNBC chemoresistance that rewires MAPK feedback and cross-talk to confer profound collateral sensitivity to combination MEK1/2 and EGFR inhibitors.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Proteínas Ativadoras de GTPase/fisiologia , Quinases de Proteína Quinase Ativadas por Mitógeno/fisiologia , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Neoplasias de Mama Triplo Negativas/genética , Linhagem Celular Tumoral , Receptores ErbB/fisiologia , Feminino , HumanosRESUMO
BACKGROUND: Metastatic melanoma is an aggressive malignancy that is resistant to therapy and has a poor prognosis. The progression of primary melanoma to metastatic disease is a multi-step process that requires dynamic regulation of gene expression through currently uncharacterized epigenetic mechanisms. Epigenetic regulation of gene expression often involves changes in chromatin structure that are catalyzed by chromatin remodeling enzymes. Understanding the mechanisms involved in the regulation of gene expression during metastasis is important for developing an effective strategy to treat metastatic melanoma. SWI/SNF enzymes are multisubunit complexes that contain either BRG1 or BRM as the catalytic subunit. We previously demonstrated that heterogeneous SWI/SNF complexes containing either BRG1 or BRM are epigenetic modulators that regulate important aspects of the melanoma phenotype and are required for melanoma tumorigenicity in vitro. RESULTS: To characterize BRG1 expression during melanoma progression, we assayed expression of BRG1 in patient derived normal skin and in melanoma specimen. BRG1 mRNA levels were significantly higher in stage IV melanomas compared to stage III tumors and to normal skin. To determine the role of BRG1 in regulating the expression of genes involved in melanoma metastasis, we expressed BRG1 in a melanoma cell line that lacks BRG1 expression and examined changes in extracellular matrix and adhesion molecule expression. We found that BRG1 modulated the expression of a subset of extracellular matrix remodeling enzymes and adhesion proteins. Furthermore, BRG1 altered melanoma adhesion to different extracellular matrix components. Expression of BRG1 in melanoma cells that lack BRG1 increased invasive ability while down-regulation of BRG1 inhibited invasive ability in vitro. Activation of metalloproteinase (MMP) 2 expression greatly contributed to the BRG1 induced increase in melanoma invasiveness. We found that BRG1 is recruited to the MMP2 promoter and directly activates expression of this metastasis associated gene. CONCLUSIONS: We provide evidence that BRG1 expression increases during melanoma progression. Our study has identified BRG1 target genes that play an important role in melanoma metastasis and we show that BRG1 promotes melanoma invasive ability in vitro. These results suggest that increased BRG1 levels promote the epigenetic changes in gene expression required for melanoma metastasis to proceed.
Assuntos
DNA Helicases/metabolismo , Melanoma/metabolismo , Melanoma/patologia , Proteínas Nucleares/metabolismo , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Fatores de Transcrição/metabolismo , Antígenos CD , Antígeno CD56/genética , Antígeno CD56/metabolismo , Caderinas/genética , Caderinas/metabolismo , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , DNA Helicases/genética , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Citometria de Fluxo , Humanos , Immunoblotting , Imuno-Histoquímica , Imunoprecipitação , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Melanoma/genética , Proteínas Nucleares/genética , Reação em Cadeia da Polimerase , Neoplasias Cutâneas/genética , Fatores de Transcrição/genética , CalininaRESUMO
Mutations in histone modifying enzymes and histone variants were identified in multiple cancers in The Cancer Genome Atlas (TCGA) studies. However, very little progress and understanding has been made in identifying the contribution of epigenetic factors in head and neck squamous cell carcinoma (HNSCC). Here, we report the identification of RUVBL1 (TIP49a), a component of the TIP60 histone modifying complex as being amplified and overexpressed in HNSCC. RUVBL1 plays a key role in incorporating histone variant H2AZ in chromatin thereby regulating transcription of key genes involved in differentiation, cancer cell proliferation and invasion. H2AZ is also overexpressed in HNSCC tumors thereby regulating RUVBL1/H2AZ dependent transcriptional programs. Patient data analysis of multiple cohorts including TCGA and single cell HNSCC data indicated RUVBL1 overexpression as a poor prognostic marker and predicts poor survival. In vitro experiments indicate a pro-proliferative role for RUVBL1/H2AZ in HNSCC cells. RUVBL1 inversely correlates with differentiation program and positively correlates with oncogenic programs, making it a key contributor to tumorigenesis and a vulnerable therapeutic target in HNSCC patients.
Assuntos
ATPases Associadas a Diversas Atividades Celulares/genética , Proteínas de Transporte/genética , Diferenciação Celular/genética , Proliferação de Células/genética , DNA Helicases/genética , Neoplasias de Cabeça e Pescoço/genética , Histonas/genética , Carcinoma de Células Escamosas de Cabeça e Pescoço/genética , ATPases Associadas a Diversas Atividades Celulares/metabolismo , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Cromatina/metabolismo , DNA Helicases/metabolismo , Bases de Dados Genéticas , Epigênese Genética , Amplificação de Genes , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias de Cabeça e Pescoço/mortalidade , Neoplasias de Cabeça e Pescoço/patologia , Histonas/metabolismo , Humanos , Invasividade Neoplásica/genética , Prognóstico , RNA Mensageiro/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/mortalidade , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Transcrição GênicaRESUMO
Women harboring heterozygous germline mutations of BRCA2 have a 50 to 80% risk of developing breast cancer, yet the pathogenesis of these cancers is poorly understood. To reveal early steps in BRCA2-associated carcinogenesis, we analyzed sorted cell populations from freshly-isolated, non-cancerous breast tissues of BRCA2 mutation carriers and matched controls. Single-cell whole-genome sequencing demonstrates that >25% of BRCA2 carrier (BRCA2mut/+ ) luminal progenitor (LP) cells exhibit sub-chromosomal copy number variations, which are rarely observed in non-carriers. Correspondingly, primary BRCA2mut/+ breast epithelia exhibit DNA damage together with attenuated replication checkpoint and apoptotic responses, and an age-associated expansion of the LP compartment. We provide evidence that these phenotypes do not require loss of the wild-type BRCA2 allele. Collectively, our findings suggest that BRCA2 haploinsufficiency and associated DNA damage precede histologic abnormalities in vivo. Using these hallmarks of cancer predisposition will yield unanticipated opportunities for improved risk assessment and prevention strategies in high-risk patients.
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Proteína BRCA2/genética , Neoplasias da Mama/genética , Predisposição Genética para Doença , Haploinsuficiência/genética , Adulto , Aneuploidia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Variações do Número de Cópias de DNA/genética , Dano ao DNA/genética , Feminino , Mutação em Linhagem Germinativa/genética , Heterozigoto , Humanos , Pessoa de Meia-Idade , Análise de Célula ÚnicaRESUMO
We show that the loss or gain of transcription factor programs that govern embryonic cell-fate specification is associated with a form of tumor plasticity characterized by the acquisition of alternative cell fates normally characteristic of adjacent organs. In human non-small cell lung cancers, downregulation of the lung lineage-specifying TF NKX2-1 is associated with tumors bearing features of various gut tissues. Loss of Nkx2-1 from murine alveolar, but not airway, epithelium results in conversion of lung cells to gastric-like cells. Superimposing oncogenic Kras activation enables further plasticity in both alveolar and airway epithelium, producing tumors that adopt midgut and hindgut fates. Conversely, coupling Nkx2-1 loss with foregut lineage-specifying SOX2 overexpression drives the formation of squamous cancers with features of esophageal differentiation. These findings demonstrate that elements of pathologic tumor plasticity mirror the normal developmental history of organs in that cancer cells acquire cell fates associated with developmentally related neighboring organs.
Assuntos
Linhagem da Célula , Neoplasias Esofágicas/patologia , Neoplasias Pulmonares/patologia , Fatores de Transcrição SOXB1/metabolismo , Neoplasias Gástricas/patologia , Fator Nuclear 1 de Tireoide/metabolismo , Adenocarcinoma Mucinoso/genética , Adenocarcinoma Mucinoso/metabolismo , Adenocarcinoma Mucinoso/patologia , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Diferenciação Celular , Plasticidade Celular , Desenvolvimento Embrionário , Endoderma/metabolismo , Endoderma/patologia , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Prognóstico , Fatores de Transcrição SOXB1/genética , Transdução de Sinais , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Taxa de Sobrevida , Fator Nuclear 1 de Tireoide/genéticaRESUMO
We sought to uncover genetic drivers of hormone receptor-positive (HR+) breast cancer, using a targeted next-generation sequencing approach for detecting expressed gene rearrangements without prior knowledge of the fusion partners. We identified intergenic fusions involving driver genes, including PIK3CA, AKT3, RAF1, and ESR1, in 14% (24/173) of unselected patients with advanced HR+ breast cancer. FISH confirmed the corresponding chromosomal rearrangements in both primary and metastatic tumors. Expression of novel kinase fusions in nontransformed cells deregulates phosphoprotein signaling, cell proliferation, and survival in three-dimensional culture, whereas expression in HR+ breast cancer models modulates estrogen-dependent growth and confers hormonal therapy resistance in vitro and in vivo Strikingly, shorter overall survival was observed in patients with rearrangement-positive versus rearrangement-negative tumors. Correspondingly, fusions were uncommon (<5%) among 300 patients presenting with primary HR+ breast cancer. Collectively, our findings identify expressed gene fusions as frequent and potentially actionable drivers in HR+ breast cancer.Significance: By using a powerful clinical molecular diagnostic assay, we identified expressed intergenic fusions as frequent contributors to treatment resistance and poor survival in advanced HR+ breast cancer. The prevalence and biological and prognostic significance of these alterations suggests that their detection may alter clinical management and bring to light new therapeutic opportunities. Cancer Discov; 8(3); 336-53. ©2017 AACR.See related commentary by Natrajan et al., p. 272See related article by Liu et al., p. 354This article is highlighted in the In This Issue feature, p. 253.
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Neoplasias da Mama/genética , Neoplasias da Mama/mortalidade , Fusão Gênica , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases/genética , Receptor alfa de Estrogênio/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Camundongos Nus , Pessoa de Meia-Idade , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-raf/genética , Piridonas/farmacologia , Pirimidinonas/farmacologia , Receptores de Esteroides/metabolismo , Proteínas Quinases S6 Ribossômicas/genética , Proteínas Quinases S6 Ribossômicas/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Loss-of-function mutations in SWI/SNF chromatin-remodeling subunit genes are observed in many cancers, but an oncogenic role for SWI/SNF is not well established. Here, we reveal that ACTL6A, encoding an SWI/SNF subunit linked to stem cell and progenitor cell function, is frequently co-amplified and highly expressed together with the p53 family member p63 in head and neck squamous cell carcinoma (HNSCC). ACTL6A and p63 physically interact, cooperatively controlling a transcriptional program that promotes proliferation and suppresses differentiation, in part through activation of the Hippo-YAP pathway via regulators including WWC1. Ectopic ACTL6A/p63 expression promotes tumorigenesis, while ACTL6A expression and YAP activation are highly correlated in primary HNSCC and predict poor patient survival. Thus, ACTL6A and p63 collaborate as oncogenic drivers in HNSCC.
Assuntos
Actinas/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Carcinoma de Células Escamosas/etiologia , Proteínas Cromossômicas não Histona/fisiologia , Proteínas de Ligação a DNA/fisiologia , Neoplasias de Cabeça e Pescoço/etiologia , Proteínas de Membrana/fisiologia , Fosfoproteínas/fisiologia , Animais , Carcinoma de Células Escamosas/mortalidade , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias de Cabeça e Pescoço/mortalidade , Via de Sinalização Hippo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Camundongos , Prognóstico , Proteínas Serina-Treonina Quinases/fisiologia , Carcinoma de Células Escamosas de Cabeça e Pescoço , Fatores de Transcrição , Transcriptoma , Proteínas de Sinalização YAPRESUMO
The mammary epithelium is organized as a bilayer of luminal and basal/myoepithelial cells. During pregnancy, the luminal compartment expands for milk production, while basal cells are thought to provide structural and contractile support. Here, we reveal a pregnancy-specific role of basal epithelia as a central coordinator of lactogenesis. We demonstrate that genetic deletion of the transcription factor p63 (Trp63) gene exclusively within basal cells of the adult gland during pregnancy leads to dramatic defects in luminal cell proliferation and differentiation, resulting in lactation failure. This phenotype is explained by direct transcriptional activation of the epidermal growth factor family ligand gene Nrg1 by p63 selectively in basal cells, which is required for luminal ERBB4/STAT5A activation and consequent luminal progenitor cell maturation. Thus, paracrine basal-to-luminal cell signaling, controlled by p63 via NRG1, orchestrates the entire lactation program. Collectively, these findings redefine the paradigm for cellular interactions specifying the functional maturation of the mammary gland.
Assuntos
Células-Tronco Adultas/metabolismo , Células Epiteliais/metabolismo , Lactação , Neuregulina-1/metabolismo , Fosfoproteínas/metabolismo , Transativadores/metabolismo , Células-Tronco Adultas/citologia , Células-Tronco Adultas/fisiologia , Animais , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células , Células Epiteliais/citologia , Células Epiteliais/fisiologia , Receptores ErbB/genética , Receptores ErbB/metabolismo , Feminino , Deleção de Genes , Células HEK293 , Humanos , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/fisiologia , Camundongos , Neuregulina-1/genética , Comunicação Parácrina , Fosfoproteínas/genética , Gravidez/metabolismo , Receptor ErbB-4 , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/metabolismo , Transativadores/genética , Ativação TranscricionalRESUMO
Our understanding of how stem cells are regulated to maintain appropriate tissue size and architecture is incomplete. We show that Yap (Yes-associated protein 1) is required for the actual maintenance of an adult mammalian stem cell. Without Yap, adult airway basal stem cells are lost through their unrestrained differentiation, resulting in the simplification of a pseudostratified epithelium into a columnar one. Conversely, Yap overexpression increases stem cell self-renewal and blocks terminal differentiation, resulting in epithelial hyperplasia and stratification. Yap overexpression in differentiated secretory cells causes them to partially reprogram and adopt a stem cell-like identity. In contrast, Yap knockdown prevents the dedifferentiation of secretory cells into stem cells. We then show that Yap functionally interacts with p63, the cardinal transcription factor associated with myriad epithelial basal stem cells. In aggregate, we show that Yap regulates all of the cardinal behaviors of airway epithelial stem cells and determines epithelial architecture.