RESUMEN
The paracrine actions of adipokine plasminogen activator inhibitor-1 (PAI-1) are implicated in obesity-associated tumorigenesis. Here, we show that PAI-1 mediates extracellular matrix (ECM) signaling via epigenetic repression of DKK1 in endometrial epithelial cells (EECs). While the loss of DKK1 is known to increase ß-catenin accumulation for WNT signaling activation, this epigenetic repression causes ß-catenin release from transmembrane integrins. Furthermore, PAI-1 elicits the disengagement of TIMP2 and SPARC from integrin-ß1 on the cell surface, lifting an integrin-ß1-ECM signaling constraint. The heightened interaction of integrin-ß1 with type 1 collagen (COL1) remodels extracellular fibrillar structures in the ECM. Consequently, the enhanced nanomechanical stiffness of this microenvironment is conducive to EEC motility and neoplastic transformation. The formation of extensively branched COL1 fibrils is also observed in endometrial tumors of patients with obesity. The findings highlight PAI-1 as a contributor to enhanced integrin-COL1 engagement and extensive ECM remodeling during obesity-associated neoplastic development.
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While macrophage phagocytosis is an immune defense mechanism against invading cellular organisms, cancer cells expressing the CD47 ligand send forward signals to repel this engulfment. Here we report that the reverse signaling using CD47 as a receptor additionally enhances a pro-survival function of prostate cancer cells under phagocytic attack. Although low CD47-expressing cancer cells still allow phagocytosis, the reverse signaling delays the process, leading to incomplete digestion of the entrapped cells and subsequent tumor hybrid cell (THC) formation. Viable THCs acquire c-Myc from parental cancer cells to upregulate both M1- and M2-like macrophage polarization genes. Consequently, THCs imitating dual macrophage features can confound immunosurveillance, gaining survival advantage in the host. Furthermore, these cells intrinsically express low levels of androgen receptor and its targets, resembling an adenocarcinoma-immune subtype of metastatic castration-resistant prostate cancer. Therefore, phagocytosis-generated THCs may represent a potential target for treating the disease.
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Antígeno CD47 , Macrófagos , Metástasis de la Neoplasia , Fagocitosis , Proteínas Proto-Oncogénicas c-myc , Escape del Tumor , Humanos , Masculino , Proteínas Portadoras , Antígeno CD47/metabolismo , Macrófagos/metabolismo , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/inmunología , Neoplasias de la Próstata/patología , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/inmunología , Transducción de Señal , Escape del Tumor/genética , Escape del Tumor/inmunología , Metástasis de la Neoplasia/genética , Metástasis de la Neoplasia/inmunología , Células Tumorales CultivadasRESUMEN
Tumor-associated macrophages (TAMs) are integral to the development of complex tumor microenvironments (TMEs) and can execute disparate cellular programs in response to extracellular cues. However, upstream signaling processes underpinning this phenotypic plasticity remain to be elucidated. Here, we report that concordant AXL-STAT3 signaling in TAMs is triggered by lung cancer cells or cancer-associated fibroblasts in the cytokine milieu. This paracrine action drives TAM differentiation toward a tumor-promoting "M2-like" phenotype with upregulation of CD163 and putative mesenchymal markers, contributing to TAM heterogeneity and diverse cellular functions. One of the upregulated markers, CD44, mediated by AXL-IL-11-pSTAT3 signaling cascade, enhances macrophage ability to interact with endothelial cells and facilitate formation of primitive vascular networks. We also found that AXL-STAT3 inhibition can impede the recruitment of TAMs in a xenograft mouse model, thereby suppressing tumor growth. These findings suggest the potential application of AXL-STAT3-related markers to quantitatively assess metastatic potential and inform therapeutic strategies in lung cancer.
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Neoplasias Pulmonares , Macrófagos Asociados a Tumores , Humanos , Animales , Ratones , Células Endoteliales , Transducción de Señal , Diferenciación Celular , Microambiente Tumoral , Línea Celular TumoralRESUMEN
The epigenome delineates lineage-specific transcriptional programs and restricts cell plasticity to prevent non-physiological cell fate transitions. Although cell diversification fosters tumor evolution and therapy resistance, upstream mechanisms that regulate the stability and plasticity of the cancer epigenome remain elusive. Here we show that 2-hydroxyglutarate (2HG) not only suppresses DNA repair but also mediates the high-plasticity chromatin landscape. A combination of single-cell epigenomics and multi-omics approaches demonstrates that 2HG disarranges otherwise well-preserved stable nucleosome positioning and promotes cell-to-cell variability. 2HG induces loss of motif accessibility to the luminal-defining transcriptional factors FOXA1, FOXP1, and GATA3 and a shift from luminal to basal-like gene expression. Breast tumors with high 2HG exhibit enhanced heterogeneity with undifferentiated epigenomic signatures linked to adverse prognosis. Further, ascorbate-2-phosphate (A2P) eradicates heterogeneity and impairs growth of high 2HG-producing breast cancer cells. These findings suggest 2HG as a key determinant of cancer plasticity and provide a rational strategy to counteract tumor cell evolution.
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Cromatina/metabolismo , Glutaratos/metabolismo , Oxidorreductasas de Alcohol/metabolismo , Ácido Ascórbico/análogos & derivados , Ácido Ascórbico/metabolismo , Diferenciación Celular , Línea Celular Tumoral , Reparación del ADN/fisiología , Epigenoma/genética , Factores de Transcripción Forkhead/genética , Expresión Génica/genética , Regulación de la Expresión Génica/genética , Humanos , Isocitrato Deshidrogenasa/genética , Neoplasias/genética , Neoplasias/metabolismo , Nucleosomas/metabolismo , Proteínas Represoras/genéticaRESUMEN
Endometriosis is a debilitating gynecologic disorder that affects â¼10% of women of reproductive age. Endometriosis is characterized by growth of endometriosis lesions within the abdominal cavity, generally thought to arise from retrograde menstruation of shed endometrial tissue. While the pathophysiology underlying peritoneal endometriosis lesion formation is still unclear, the interaction between invading endometrial tissue and the peritoneal mesothelial lining is an essential step in lesion formation. In this study, we assessed proteomic differences between eutopic endometrial stromal cells (ESCs) from women with and without endometriosis in response to peritoneal mesothelial cell (PMC) exposure, using single-cell cytometry by time-of-flight (CyTOF). Co-cultured primary eutopic ESCs from women with and without endometriosis with an established PMC line were subjected to immunostaining with a panel of Maxpar CyTOF metal-conjugated antibodies (n = 28) targeting cell junction and mesenchymal markers, which are involved in cell-cell adhesions and epithelial-mesenchymal transition. Exposure of the ESCs to PMCs resulted in a drastic shift in cellular expression profiles in ESCs derived from endometriosis, whereas little effect by PMCs was observed in ESCs from non-endometriosis subjects. The transcription factor SNAI1 was consistently repressed by PMC interactions. ESCs from endometriosis patients are unique in that they respond to PMCs by undergoing changes in adhesive properties and mesenchymal characteristics that would facilitate lesion formation.
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Biomarcadores/metabolismo , Endometriosis/metabolismo , Endometrio/citología , Epitelio/metabolismo , Uniones Intercelulares/metabolismo , Proteómica/métodos , Células Cultivadas , Técnicas de Cocultivo , Biología Computacional , Endometrio/metabolismo , Endometrio/patología , Células Epiteliales/citología , Células Epiteliales/metabolismo , Femenino , Humanos , Análisis de la Célula Individual , Células del Estroma/citología , Células del Estroma/metabolismoRESUMEN
While plasminogen activator inhibitor-1 (PAI-1) is known to potentiate cellular migration via proteolytic regulation, this adipokine is implicated as an oncogenic ligand in the tumor microenvironment. To understand the underlying paracrine mechanism, here, we conduct transcriptomic analysis of 1,898 endometrial epithelial cells (EECs) exposed and unexposed to PAI-1-secreting adipose stromal cells. The PAI-1-dependent action deregulates crosstalk among tumor-promoting and tumor-repressing pathways, including transforming growth factor ß (TGF-ß). When PAI-1 is tethered to lipoprotein receptor-related protein 1 (LRP1), the internalized signaling causes downregulation of SMAD4 at the transcriptional and post-translational levels that attenuates TGF-ß-related transcription programs. Repression of genes encoding the junction and adhesion complex preferentially occurs in SMAD4-underexpressed EECs of persons with obesity. The findings highlight a role of PAI-1 signaling that renders ineffective intercellular communication for the development of adiposity-associated endometrial cancer.
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Neoplasias Endometriales/metabolismo , Moléculas de Adhesión de Unión/metabolismo , Obesidad/metabolismo , Inhibidor 1 de Activador Plasminogénico/metabolismo , Proteína Smad4/metabolismo , Tejido Adiposo/patología , Regulación hacia Abajo/genética , Neoplasias Endometriales/complicaciones , Neoplasias Endometriales/genética , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Proteína 1 Relacionada con Receptor de Lipoproteína de Baja Densidad/metabolismo , Obesidad/complicaciones , Unión Proteica , Proteolisis , Proteómica , Proteína Smad4/genética , Células del Estroma/metabolismo , Transcripción Genética , Factor de Crecimiento Transformador beta/metabolismo , Microambiente Tumoral , Ubiquitina/metabolismoRESUMEN
BACKGROUND: Chromothripsis is an event of genomic instability leading to complex chromosomal alterations in cancer. Frequent long-range chromatin interactions between transcription factors (TFs) and targets may promote extensive translocations and copy-number alterations in proximal contact regions through inappropriate DNA stitching. Although studies have proposed models to explain the initiation of chromothripsis, few discussed how TFs influence this process for tumor progression. METHODS: This study focused on genomic alterations in amplification associated regions within chromosome 17. Inter-/intra-chromosomal rearrangements were analyzed using whole genome sequencing data of breast tumors in the Cancer Genome Atlas (TCGA) cohort. Common ERα binding sites were defined based on MCF-7, T47D, and MDA-MB-134 breast cancer cell lines using univariate K-means clustering methods. Nanopore sequencing technology was applied to validate frequent rearrangements detected between ATC loci on 17q23 and an ERα hub on 20q13. The efficacy of pharmacological inhibition of a potentially druggable target gene on 17q23 was evaluated using breast cancer cell lines and patient-derived circulating breast tumor cells. RESULTS: There are five adjoining regions from 17q11.1 to 17q24.1 being hotspots of chromothripsis. Inter-/intra-chromosomal rearrangements of these regions occurred more frequently in ERα-positive tumors than in ERα-negative tumors. In addition, the locations of the rearrangements were often mapped within or close to dense ERα binding sites localized on these five 17q regions or other chromosomes. This chromothriptic event was linked to concordant upregulation of 96 loci that predominantly regulate cell-cycle machineries in advanced luminal tumors. Genome-editing analysis confirmed that an ERα hub localized on 20q13 coordinately regulates a subset of these loci localized on 17q23 through long-range chromosome interactions. One of these loci, Tousled Like Kinase 2 (TLK2) known to participate in DNA damage checkpoint control, is an actionable target using phenothiazine antipsychotics (PTZs). The antiproliferative effect of PTZs was prominent in high TLK2-expressing cells, compared to low expressing cells. CONCLUSION: This study demonstrates a new approach for identifying tumorigenic drivers from genomic regions highly susceptible to ERα-related chromothripsis. We found a group of luminal breast tumors displaying 17q-related chromothripsis for which antipsychotics can be repurposed as treatment adjuncts.
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Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/genética , Cromosomas Humanos Par 17/genética , Cromotripsis , Receptor alfa de Estrógeno/metabolismo , Regulación Neoplásica de la Expresión Génica , Biomarcadores de Tumor/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ciclo Celular , Proliferación Celular , Receptor alfa de Estrógeno/genética , Femenino , Humanos , Pronóstico , Tasa de Supervivencia , Transcripción Genética , Células Tumorales Cultivadas , Secuenciación del Exoma , Secuenciación Completa del GenomaRESUMEN
SULT2B1b (SULT2B) is a prostate-expressed hydroxysteroid sulfotransferase, which may regulate intracrine androgen homeostasis by mediating 3ß-sulfation of dehydroepiandrosterone (DHEA), the precursor for 5α-dihydrotestosterone (DHT) biosynthesis. The aldo-keto reductase (AKR)1C3 regulates androgen receptor (AR) activity in castration-resistant prostate cancer (CRPC) by promoting tumor tissue androgen biosynthesis from adrenal DHEA and also by functioning as an AR-selective coactivator. Herein we report that SULT2B-depleted CRPC cells, arising from stable RNA interference or gene knockout (KO), are markedly upregulated for AKR1C3, activated for ERK1/2 survival signal, and induced for epithelial-to-mesenchymal (EMT)-like changes. EMT was evident from increased mesenchymal proteins and elevated EMT-inducing transcription factors SNAI1 and TWIST1 in immunoblot and single-cell mass cytometry analyses. SULT2B KO cells showed greater motility and invasion in vitro; growth escalation in xenograft study; and enhanced metastatic potential predicted on the basis of decreased cell stiffness and adhesion revealed from atomic force microscopy analysis. While AR and androgen levels were unchanged, AR activity was elevated, since PSA and FKBP5 mRNA induction by DHT-activated AR was several-fold higher in SULT2B-silenced cells. AKR1C3 silencing prevented ERK1/2 activation and SNAI1 induction in SULT2B-depleted cells. SULT2B was undetectable in nearly all CRPC metastases from 50 autopsy cases. Primary tumors showed variable and Gleason score (GS)-independent SULT2B levels. CRPC metastases lacking SULT2B expressed AKR1C3. Since AKR1C3 is frequently elevated in advanced prostate cancer, the inhibitory influence of SULT2B on AKR1C3 upregulation, ERK1/2 activation, EMT-like induction, and on cell motility and invasiveness may be clinically significant. Pathways regulating the inhibitory SULT2B-AKR1C3 axis may inform new avenue(s) for targeting SULT2B-deficient prostate cancer.
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Miembro C3 de la Familia 1 de las Aldo-Ceto Reductasas/metabolismo , Carcinoma/enzimología , Neoplasias de la Próstata/enzimología , Sulfotransferasas/metabolismo , Animales , Transición Epitelial-Mesenquimal , Humanos , Masculino , Ratones Desnudos , Metástasis de la Neoplasia , Trasplante de Neoplasias , Receptores Androgénicos/metabolismoRESUMEN
Cytometry by time-of-flight (CyTOF) simultaneously measures multiple cellular proteins at the single-cell level and is used to assess intertumor and intratumor heterogeneity. This approach may be used to investigate the variability of individual tumor responses to treatments. Herein, we stratified lung tumor subpopulations based on AXL signaling as a potential targeting strategy. Integrative transcriptome analyses were used to investigate how TP-0903, an AXL kinase inhibitor, influences redundant oncogenic pathways in metastatic lung cancer cells. CyTOF profiling revealed that AXL inhibition suppressed SMAD4/TGFß signaling and induced JAK1-STAT3 signaling to compensate for the loss of AXL. Interestingly, high JAK1-STAT3 was associated with increased levels of AXL in treatment-naïve tumors. Tumors with high AXL, TGFß, and JAK1 signaling concomitantly displayed CD133-mediated cancer stemness and hybrid epithelial-to-mesenchymal transition features in advanced-stage patients, suggesting greater potential for distant dissemination. Diffusion pseudotime analysis revealed cell-fate trajectories among four different categories that were linked to clinicopathologic features for each patient. Patient-derived organoids (PDO) obtained from tumors with high AXL and JAK1 were sensitive to TP-0903 and ruxolitinib (JAK inhibitor) treatments, supporting the CyTOF findings. This study shows that single-cell proteomic profiling of treatment-naïve lung tumors, coupled with ex vivo testing of PDOs, identifies continuous AXL, TGFß, and JAK1-STAT3 signal activation in select tumors that may be targeted by combined AXL-JAK1 inhibition. SIGNIFICANCE: Single-cell proteomic profiling of clinical samples may facilitate the optimal selection of novel drug targets, interpretation of early-phase clinical trial data, and development of predictive biomarkers valuable for patient stratification.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Janus Quinasa 1/antagonistas & inhibidores , Neoplasias Pulmonares/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Anciano , Anciano de 80 o más Años , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Línea Celular Tumoral , Resistencia a Antineoplásicos , Sinergismo Farmacológico , Transición Epitelial-Mesenquimal/efectos de los fármacos , Estudios de Factibilidad , Femenino , Citometría de Flujo/métodos , Humanos , Janus Quinasa 1/metabolismo , Pulmón/patología , Neoplasias Pulmonares/patología , Masculino , Ratones , Persona de Mediana Edad , Nitrilos , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteómica/métodos , Proteínas Proto-Oncogénicas/metabolismo , Pirazoles/farmacología , Pirazoles/uso terapéutico , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , RNA-Seq , Proteínas Tirosina Quinasas Receptoras/metabolismo , Transducción de Señal/efectos de los fármacos , Análisis de la Célula Individual/métodos , Sulfonamidas/farmacología , Sulfonamidas/uso terapéutico , Análisis de Matrices Tisulares , Ensayos Antitumor por Modelo de Xenoinjerto , Tirosina Quinasa del Receptor AxlRESUMEN
Advanced prostate cancer is a very heterogeneous disease reflecting in diverse regulations of oncogenic signaling pathways. Aberrant spatial dynamics of epidermal growth factor receptor (EGFR) promote their dimerization and clustering, leading to constitutive activation in oncogenesis. The EphB2 and Src signaling pathways are associated with the reorganization of the cytoskeleton leading to malignancy, but their roles in regulating EGFR dynamics and activation are scarcely reported. Using single-particle tracking techniques, we found that highly phosphorylated EGFR in the advanced prostate cancer cell line, PC3, was associated with higher EGFR diffusivity, as compared with LNCaP and less aggressive DU145. The increased EGFR activation and biophysical dynamics were consistent with high proliferation, migration, and invasion. After performing single-cell RNA-seq on prostate cancer cell lines and circulating tumor cells from patients, we identified that upregulated gene expression in the EphB2 and Src pathways are associated with advanced malignancy. After dasatinib treatment or siRNA knockdowns of EphB2 or Src, the PC3 cells exhibited significantly lower EGFR dynamics, cell motility, and invasion. Partial inhibitory effects were also found in DU145 cells. The upregulation of parts of the EphB2 and Src pathways also predicts poor prognosis in the prostate cancer patient cohort of The Cancer Genome Atlas. Our results provide evidence that overexpression of the EphB2 and Src signaling pathways regulate EGFR dynamics and cellular aggressiveness in some advanced prostate cancer cells.
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Hepatocellular carcinomas (HCC) exhibit distinct promoter hypermethylation patterns, but the epigenetic regulation and function of transcriptional enhancers remain unclear. Here, our affinity- and bisulfite-based whole-genome sequencing analyses reveal global enhancer hypomethylation in human HCCs. Integrative epigenomic characterization further pinpoints a recurrent hypomethylated enhancer of CCAAT/enhancer-binding protein-beta (C/EBPß) which correlates with C/EBPß over-expression and poorer prognosis of patients. Demethylation of C/EBPß enhancer reactivates a self-reinforcing enhancer-target loop via direct transcriptional up-regulation of enhancer RNA. Conversely, deletion of this enhancer via CRISPR/Cas9 reduces C/EBPß expression and its genome-wide co-occupancy with BRD4 at H3K27ac-marked enhancers and super-enhancers, leading to drastic suppression of driver oncogenes and HCC tumorigenicity. Hepatitis B X protein transgenic mouse model of HCC recapitulates this paradigm, as C/ebpß enhancer hypomethylation associates with oncogenic activation in early tumorigenesis. These results support a causal link between aberrant enhancer hypomethylation and C/EBPß over-expression, thereby contributing to hepatocarcinogenesis through global transcriptional reprogramming.
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Carcinogénesis/genética , Metilación de ADN , Neoplasias Hepáticas/genética , Secuencias Reguladoras de Ácidos Nucleicos/genética , Animales , Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Carcinoma Hepatocelular/genética , Proteínas de Ciclo Celular , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Desmetilación , Epigénesis Genética , Eliminación de Gen , Regulación Neoplásica de la Expresión Génica , Humanos , Hígado , Ratones , Ratones Transgénicos , Proteínas Nucleares/metabolismo , Pronóstico , Regiones Promotoras Genéticas , Transactivadores , Factores de Transcripción/metabolismo , Activación Transcripcional , Regulación hacia Arriba , Proteínas Reguladoras y Accesorias ViralesRESUMEN
Emerging evidence indicates that adipose stromal cells (ASC) are recruited to enhance cancer development. In this study, we examined the role these adipocyte progenitors play relating to intercellular communication in obesity-associated endometrial cancer. This is particularly relevant given that gap junctions have been implicated in tumor suppression. Examining the effects of ASCs on the transcriptome of endometrial epithelial cells (EEC) in an in vitro coculture system revealed transcriptional repression of GJA1 (encoding the gap junction protein Cx43) and other genes related to intercellular communication. This repression was recapitulated in an obesity mouse model of endometrial cancer. Furthermore, inhibition of plasminogen activator inhibitor 1 (PAI-1), which was the most abundant ASC adipokine, led to reversal of cellular distribution associated with the GJA1 repression profile, suggesting that PAI-1 may mediate actions of ASC on transcriptional regulation in EEC. In an endometrial cancer cohort (n = 141), DNA hypermethylation of GJA1 and related loci TJP2 and PRKCA was observed in primary endometrial endometrioid tumors and was associated with obesity. Pharmacologic reversal of DNA methylation enhanced gap-junction intercellular communication and cell-cell interactions in vitro. Restoring Cx43 expression in endometrial cancer cells reduced cellular migration; conversely, depletion of Cx43 increased cell migration in immortalized normal EEC. Our data suggest that persistent repression by ASC adipokines leads to promoter hypermethylation of GJA1 and related genes in the endometrium, triggering long-term silencing of these loci in endometrial tumors of obese patients. SIGNIFICANCE: Studies reveal that adipose-derived stem cells in endometrial cancer pathogenesis influence epigenetic repression of gap junction loci, which suggests targeting of gap junction activity as a preventive strategy for obesity-associated endometrial cancer.
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Adipoquinas/farmacología , Tejido Adiposo/patología , Comunicación Celular , Conexina 43/genética , Neoplasias Endometriales/patología , Represión Epigenética , Obesidad/complicaciones , Tejido Adiposo/metabolismo , Animales , Movimiento Celular , Células Cultivadas , Conexina 43/metabolismo , Dieta Alta en Grasa/efectos adversos , Neoplasias Endometriales/etiología , Neoplasias Endometriales/metabolismo , Células Epiteliales/metabolismo , Células Epiteliales/patología , Femenino , Uniones Comunicantes , Humanos , Masculino , Ratones , Ratones Noqueados , Obesidad/fisiopatología , Células del Estroma/metabolismo , Células del Estroma/patologíaRESUMEN
Increasing evidence suggests the presence of minor cell subpopulations in prostate cancer that are androgen independent and poised for selection as dominant clones after androgen deprivation therapy. In this study, we investigated this phenomenon by stratifying cell subpopulations based on transcriptome profiling of 144 single LNCaP prostate cancer cells treated or untreated with androgen after cell-cycle synchronization. Model-based clustering of 397 differentially expressed genes identified eight potential subpopulations of LNCaP cells, revealing a previously unappreciable level of cellular heterogeneity to androgen stimulation. One subpopulation displayed stem-like features with a slower cell doubling rate, increased sphere formation capability, and resistance to G2-M arrest induced by a mitosis inhibitor. Advanced growth of this subpopulation was associated with enhanced expression of 10 cell-cycle-related genes (CCNB2, DLGAP5, CENPF, CENPE, MKI67, PTTG1, CDC20, PLK1, HMMR, and CCNB1) and decreased dependence upon androgen receptor signaling. In silico analysis of RNA-seq data from The Cancer Genome Atlas further demonstrated that concordant upregulation of these genes was linked to recurrent prostate cancers. Analysis of receiver operating characteristic curves implicates aberrant expression of these genes and could be useful for early identification of tumors that subsequently develop biochemical recurrence. Moreover, this single-cell approach provides a better understanding of how prostate cancer cells respond heterogeneously to androgen deprivation therapies and reveals characteristics of subpopulations resistant to this treatment.Significance: Illustrating the challenge in treating cancers with targeted drugs, which by selecting for drug resistance can drive metastatic progression, this study characterized the plasticity and heterogeneity of prostate cancer cells with regard to androgen dependence, defining the character or minor subpopulations of androgen-independent cells that are poised for clonal selection after androgen-deprivation therapy. Cancer Res; 78(4); 853-64. ©2017 AACR.
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Andrógenos/metabolismo , Perfilación de la Expresión Génica/métodos , Neoplasias de la Próstata/genética , ARN/metabolismo , Línea Celular Tumoral , Humanos , Masculino , Neoplasias de la Próstata/patologíaRESUMEN
Despite numerous studies done on understanding the role of DNA methylation, limited work has focused on systems integration of cell type-specific interplay between DNA methylation and gene transcription. Through a genome-wide analysis of DNA methylation across 19 cell types with T-47D as reference, we identified 106,252 cell type-specific differentially-methylated CpGs categorized into 7,537 differentially (46.6% hyper- and 53.4% hypo-) methylated regions. We found 44% promoter regions and 75% CpG islands were T-47D cell type-specific methylated. Pyrosequencing experiments validated the cell type-specific methylation across three benchmark cell lines. Interestingly, these DMRs overlapped with 1,145 known tumor suppressor genes. We then developed a Bayesian Gaussian Regression model to measure the relationship among DNA methylation, genomic segment distribution, differential gene expression and tumor suppressor gene status. The model uncovered that 3'UTR methylation has much less impact on transcriptional activity than other regions. Integration of DNA methylation and 82 transcription factor binding information across the 19 cell types suggested diverse interplay patterns between the two regulators. Our integrative analysis reveals cell type-specific and genomic region-dependent regulatory patterns and provides a perspective for integrating hundreds of various omics-seq data together.
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Metilación de ADN , Regulación de la Expresión Génica , Genómica , Transcripción Genética , Línea Celular , Línea Celular Tumoral , Islas de CpG , Perfilación de la Expresión Génica , Genómica/métodos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Especificidad de Órganos , Secuencias Reguladoras de Ácidos NucleicosRESUMEN
BACKGROUND: The Pap smear has remained the foundation for cervical cancer screening for over 70 years. With advancements in molecular diagnostics, primary high-risk human papillomavirus (hrHPV) screening has recently become an accepted stand-alone or co-test with conventional cytology. However, both diagnostic tests have distinct limitations. The aim of this study was to determine the association between HPV genotypes and cellular epigenetic modifications in three grades of cervical cytology for screening biomarker discovery. METHODS: This prospective, cross-sectional study used residual liquid-based cytology samples for HPV genotyping and epigenetic analysis. Extracted DNA was subjected to parallel polymerase chain reactions using three primer sets (MY09/11, FAP59/64, E6-E7 F/B) for HPV DNA amplification. HPV+ samples were genotyped by DNA sequencing. Promoter methylation of four candidate tumor suppressor genes (adenylate cyclase 8 (ADCY8), cadherin 8, type 2 (CDH8), MGMT, and zinc finger protein 582 (ZNF582)) out of 48 genes screened was quantified by bisulfite-pyrosequencing of genomic DNA. Independent validation of methylation profiles was performed by analyzing data from cervical cancer cell lines and clinical samples from The Cancer Genome Atlas (TCGA). RESULTS: Two hundred seventy-seven quality cytology samples were analyzed. HPV was detected in 31/100 (31 %) negative for intraepithelial lesion or malignancy (NILM), 95/100 (95 %) low-grade squamous intraepithelial lesion (LSIL), and 71/77 (92 %) high-grade squamous intraepithelial lesion (HSIL) samples. The proportion of IARC-defined carcinogenic HPV types in sequenced samples correlated with worsening grade: NILM 7/29 (24 %), LSIL 53/92 (58 %), and HSIL 65/70 (93 %). Promoter methylation of ADCY8, CDH8, and ZNF582 was measured in 170 samples: NILM (N = 33), LSIL (N = 70), and HSIL (N = 67) also correlated with worsening grade. Similar hypermethylation patterns were found in cancer cell lines and TCGA samples. The combination of four biomarkers, i.e., HPV genotype and three-gene promoter methylation, predicted HSIL (AUC 0.89) better than HPV alone (AUC 0.74) by logistic regression and probabilistic modeling. CONCLUSIONS: HPV genotype and DNA methylation of ADCY8, CDH8, and ZNF582 are correlated with cytological grade. Collectively, these biomarkers may serve as a molecular classifier of Pap smears.
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Adenilil Ciclasas/genética , Cadherinas/genética , Metilación de ADN , Factores de Transcripción de Tipo Kruppel/genética , Papillomaviridae/genética , Infecciones por Papillomavirus/diagnóstico , Neoplasias del Cuello Uterino/epidemiología , Neoplasias del Cuello Uterino/patología , Adulto , Línea Celular Tumoral , Estudios Transversales , ADN Viral/análisis , Detección Precoz del Cáncer , Femenino , Genotipo , Humanos , Persona de Mediana Edad , Clasificación del Tumor , Prueba de Papanicolaou/métodos , Infecciones por Papillomavirus/epidemiología , Regiones Promotoras Genéticas , Estudios Prospectivos , Análisis de Secuencia de ADN/métodos , Lesiones Intraepiteliales Escamosas de Cuello Uterino/epidemiología , Lesiones Intraepiteliales Escamosas de Cuello Uterino/genética , Lesiones Intraepiteliales Escamosas de Cuello Uterino/patología , Lesiones Intraepiteliales Escamosas de Cuello Uterino/virología , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/virología , Frotis Vaginal/métodos , Adulto Joven , Displasia del Cuello del Útero/epidemiología , Displasia del Cuello del Útero/genética , Displasia del Cuello del Útero/patología , Displasia del Cuello del Útero/virologíaRESUMEN
Identifying prostate cancer-driving transcription factors (TFs) in addition to the androgen receptor promises to improve our ability to effectively diagnose and treat this disease. We employed an integrative genomics analysis of master TFs CREB1 and FoxA1 in androgen-dependent prostate cancer (ADPC) and castration-resistant prostate cancer (CRPC) cell lines, primary prostate cancer tissues and circulating tumor cells (CTCs) to investigate their role in defining prostate cancer gene expression profiles. Combining genome-wide binding site and gene expression profiles we define CREB1 as a critical driver of pro-survival, cell cycle and metabolic transcription programs. We show that CREB1 and FoxA1 co-localize and mutually influence each other's binding to define disease-driving transcription profiles associated with advanced prostate cancer. Gene expression analysis in human prostate cancer samples found that CREB1/FoxA1 target gene panels predict prostate cancer recurrence. Finally, we showed that this signaling pathway is sensitive to compounds that inhibit the transcription co-regulatory factor MED1. These findings not only reveal a novel, global transcriptional co-regulatory function of CREB1 and FoxA1, but also suggest CREB1/FoxA1 signaling is a targetable driver of prostate cancer progression and serves as a biomarker of poor clinical outcomes.
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Proteína de Unión a Elemento de Respuesta al AMP Cíclico/fisiología , Factor Nuclear 3-alfa del Hepatocito/fisiología , Recurrencia Local de Neoplasia/metabolismo , Neoplasias de la Próstata/metabolismo , Anciano , Anciano de 80 o más Años , Secuencia de Bases , Sitios de Unión , Biomarcadores de Tumor , Línea Celular Tumoral , Secuencia de Consenso , Supervivencia sin Enfermedad , Regulación Neoplásica de la Expresión Génica , Ontología de Genes , Humanos , Estimación de Kaplan-Meier , Masculino , Subunidad 1 del Complejo Mediador/metabolismo , Persona de Mediana Edad , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/mortalidad , Pronóstico , Modelos de Riesgos Proporcionales , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/mortalidad , Neoplasias de la Próstata/patología , Inhibidores de Proteínas Quinasas/farmacología , Transducción de Señal , Transcripción GenéticaRESUMEN
BACKGROUND: Altered DNA methylation in CpG islands of gene promoters has been implicated in prostate cancer (PCa) progression and can be used to predict disease outcome. In this study, we determine whether methylation changes of androgen biosynthesis pathway (ABP)-related genes in patients' plasma cell-free DNA (cfDNA) can serve as prognostic markers for biochemical recurrence (BCR). METHODS: Methyl-binding domain capture sequencing (MBDCap-seq) was used to identify differentially methylated regions (DMRs) in primary tumors of patients who subsequently developed BCR or not, respectively. Methylation pyrosequencing of candidate loci was validated in cfDNA samples of 86 PCa patients taken at and/or post-radical prostatectomy (RP) using univariate and multivariate prediction analyses. RESULTS: Putative DMRs in 13 of 30 ABP-related genes were found between tumors of BCR (n = 12) versus no evidence of disease (NED) (n = 15). In silico analysis of The Cancer Genome Atlas data confirmed increased DNA methylation of two loci-SRD5A2 and CYP11A1, which also correlated with their decreased expression, in tumors with subsequent BCR development. Their aberrant cfDNA methylation was also associated with detectable levels of PSA taken after patients' post-RP. Multivariate analysis of the change in cfDNA methylation at all of CpG sites measured along with patient's treatment history predicted if a patient will develop BCR with 77.5% overall accuracy. CONCLUSIONS: Overall, increased DNA methylation of SRD5A2 and CYP11A1 related to androgen biosynthesis functions may play a role in BCR after patients' RP. The correlation between aberrant cfDNA methylation and detectable PSA in post-RP further suggests their utility as predictive markers for PCa recurrence. .
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3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/genética , Enzima de Desdoblamiento de la Cadena Lateral del Colesterol/genética , Metilación de ADN , Proteínas de la Membrana/genética , Recurrencia Local de Neoplasia/genética , Neoplasias de la Próstata/genética , Anciano , Biomarcadores de Tumor/genética , Islas de CpG , Supervivencia sin Enfermedad , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/patología , Regiones Promotoras Genéticas , Próstata/patología , Próstata/cirugía , Prostatectomía , Neoplasias de la Próstata/patología , Neoplasias de la Próstata/cirugía , Factores de RiesgoRESUMEN
We examined epigenetic regulation in regards to behaviorally and clinically relevant human brain function. Specifically, we found that increased promoter methylation of the serotonin transporter gene predicted increased threat-related amygdala reactivity and decreased mRNA expression in postmortem amygdala tissue. These patterns were independent of functional genetic variation in the same region. Furthermore, the association with amygdala reactivity was replicated in a second cohort and was robust to both sampling methods and age.
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Amígdala del Cerebelo/fisiología , Epigénesis Genética/fisiología , Imagen por Resonancia Magnética , Modelos Neurológicos , Proteínas de Transporte de Serotonina en la Membrana Plasmática/genética , Proteínas de Transporte de Serotonina en la Membrana Plasmática/fisiología , Adolescente , Niño , Metilación de ADN/fisiología , Femenino , Genotipo , Humanos , Modelos Lineales , Masculino , Valor Predictivo de las Pruebas , Regiones Promotoras Genéticas/fisiología , Adulto JovenRESUMEN
BACKGROUND: Emerging evidence shows that nanomechanical phenotypes of circulating tumor cells (CTC) could become potential biomarkers for metastatic castration resistant prostate cancer (mCRPC). METHODS: To determine the nanomechanical phenotypes of CTCs we applied atomic force microscopy (AFM) employing the PeakForce quantitative nanomechanical (QNM) imaging. We assessed biophysical parameters (elasticity, deformation, and adhesion) of 130 CTCs isolated from blood samples from five castration sensitive (CS) and 12 castration resistant prostate cancer (CRPCa) patients. RESULTS: We found that CTCs from CRPCa patients are three times softer, three times more deformable, and seven times more adhesive than counterparts from CSPCa patients. Both nonsupervised hierarchical clustering and principle component analysis show that three combined nanomechanical parameters could constitute a valuable set to distinguish between CSPCa and CRPCa. CONCLUSIONS: [corrected] Our study indicates that nanomechanical phenotypes of CTCs may serve as novel and effective biomarkers for mCRPC.