RESUMO
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.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Fator 3-alfa Nuclear de Hepatócito/fisiologia , Recidiva Local de Neoplasia/metabolismo , Neoplasias da Próstata/metabolismo , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Sítios de Ligação , Biomarcadores Tumorais , Linhagem Celular Tumoral , Sequência Consenso , Intervalo Livre de Doença , Regulação Neoplásica da Expressão Gênica , Ontologia Genética , Humanos , Estimativa de Kaplan-Meier , Masculino , Subunidade 1 do Complexo Mediador/metabolismo , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/mortalidade , Prognóstico , Modelos de Riscos Proporcionais , Neoplasias da Próstata/genética , Neoplasias da Próstata/mortalidade , Neoplasias da Próstata/patologia , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais , Transcrição GênicaRESUMO
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. .
Assuntos
3-Oxo-5-alfa-Esteroide 4-Desidrogenase/genética , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Metilação de DNA , Proteínas de Membrana/genética , Recidiva Local de Neoplasia/genética , Neoplasias da Próstata/genética , Idoso , Biomarcadores Tumorais/genética , Ilhas de CpG , Intervalo Livre de Doença , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Regiões Promotoras Genéticas , Próstata/patologia , Próstata/cirurgia , Prostatectomia , Neoplasias da Próstata/patologia , Neoplasias da Próstata/cirurgia , Fatores de RiscoRESUMO
Recent clinical and pre-clinical studies suggest that both active and passive immunization strategies targeting Aß amyloid may have clinical benefit in Alzheimer's disease. Here, we demonstrate that vaccination of APPswePSEN1dE9 mice with SDPM1, an engineered non-native Aß amyloid-specific binding peptide, lowers brain Aß amyloid plaque burden and brain Aß1-40 and Aß1-42 peptide levels, improves cognitive learning and memory in Morris water maze tests and increases the expression of synaptic brain proteins. This was the case in young mice immunized prior to development of significant brain amyloid burden, and in older mice, where brain amyloid was already present. Active immunization was optimized using ALUM as an adjuvant to stimulate production of anti-SDPM1 and anti-Aß amyloid antibodies. Intracerebral injection of P4D6, an SDPM1 peptide-mimotope antibody, also lowered brain amyloid plaque burden in APPswePSEN1dE9 mice. Additionally, P4D6 inhibited Aß amyloid-mediated toxicity in cultured neuronal cells. The protein sequence of the variable domain within the P4D6 heavy chain was found to mimic a multimer of the SDPM1 peptide motif. These data demonstrate the efficacy of active and passive vaccine strategies to target Aß amyloid oligomers using an engineered peptide-mimotope strategy.
Assuntos
Doença de Alzheimer/terapia , Vacinas contra Alzheimer/uso terapêutico , Peptídeos/uso terapêutico , Hidróxido de Alumínio/imunologia , Doença de Alzheimer/patologia , Doença de Alzheimer/prevenção & controle , Peptídeos beta-Amiloides/metabolismo , Animais , Córtex Cerebral/patologia , Modelos Animais de Doenças , Hipocampo/patologia , Imunização Passiva , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Camundongos Transgênicos , Fragmentos de Peptídeos/metabolismo , Placa Amiloide/patologia , Sinapses/metabolismo , Resultado do Tratamento , VacinaçãoRESUMO
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.
Assuntos
Biomarcadores Tumorais/sangue , Células Neoplásicas Circulantes/patologia , Neoplasias de Próstata Resistentes à Castração/diagnóstico , Neoplasias da Próstata/diagnóstico , Contagem de Células , Humanos , Masculino , Células Neoplásicas Circulantes/metabolismo , Neoplasias da Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/sangue , Neoplasias de Próstata Resistentes à Castração/patologiaRESUMO
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.
RESUMO
DNA hypermethylation of promoter CpG islands is associated with epigenetic silencing of tumor suppressor genes in oral squamous cell carcinomas (OSCCs). We used a methyl-CpG-binding domain protein capture method coupled with next-generation sequencing (MBDCap-seq) to survey global DNA methylation patterns in OSCCs with and without nodal metastasis and normal mucosa (total n = 58). Of 1462 differentially methylated CpG islands identified in OSCCs relative to normal controls, MBDCap-seq profiling uncovered 359 loci linked to lymph node metastasis. Interactive network analysis revealed a subset of these loci (n = 23), including the anaplastic lymphoma kinase (ALK) gene, are potential regulators and effectors of invasiveness and metastatic progression. Promoter methylation of ALK was preferentially observed in OSCCs without node metastasis, whereas relatively lower methylation levels were present in metastatic tumors, implicating an active state of ALK transcription in the latter group. The OSCC cell line, SCC4, displayed reduced ALK expression that corresponded to extensive promoter CpG island methylation. SCC4 treatment with demethylating agents induced ALK expression and increased invasion and migration characteristics. Inhibition of ALK activity in OSCC cells with high ALK expression (CAL27, HSC3 and SCC25), decreased cell growth and resulted in changes in invasive potential and mesenchymal marker expression that were cell-line dependent. Although ALK is susceptible to epigenetic silencing during oral tumorigenesis, overwriting this default state may be necessary for modulating invasive processes involved in nodal metastases. Given the complex response of OSCC cells to ALK inhibition, future studies are required to assess the feasibility of targeting ALK to treat invasive OSCCs.
Assuntos
Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Mesoderma/patologia , Neoplasias Bucais/genética , Neoplasias Bucais/patologia , Receptores Proteína Tirosina Quinases/genética , Quinase do Linfoma Anaplásico , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Carcinoma de Células Escamosas/metabolismo , Processos de Crescimento Celular/fisiologia , Linhagem Celular Tumoral , Ilhas de CpG , Metilação de DNA , Progressão da Doença , Epigênese Genética , Humanos , Metástase Linfática , Mesoderma/metabolismo , Mucosa Bucal/metabolismo , Mucosa Bucal/patologia , Neoplasias Bucais/metabolismo , Invasividade Neoplásica , Regiões Promotoras Genéticas , Receptores Proteína Tirosina Quinases/metabolismo , Ativação TranscricionalRESUMO
Roughly 3 million years ago, an inactivating deletion occurred in CMAH, the human gene encoding CMP-Neu5Ac (cytidine-5'-monophospho-N-acetylneuraminic acid) hydroxylase (Chou HH, Takematsu H, Diaz S, Iber J, Nickerson E, Wright KL, Muchmore EA, Nelson DL, Warren ST, Varki A. 1998. A mutation in human CMP-sialic acid hydroxylase occurred after the Homo-Pan divergence. Proc Natl Acad Sci USA. 95:11751-11756). This inactivating deletion is now homozygous in all humans, causing the loss of N-glycolylneuraminic acid (Neu5Gc) biosynthesis in all human cells and tissues. The CMAH enzyme is active in other mammals, including mice, where Neu5Gc is an abundant form of sialic acid on cellular membranes, including those in cardiac and skeletal muscle. We recently demonstrated that the deletion of mouse Cmah worsened the severity of pathophysiology measures related to muscular dystrophy in mdx mice, a model for Duchenne muscular dystrophy (Chandrasekharan K, Yoon JH, Xu Y, deVries S, Camboni M, Janssen PM, Varki A, Martin PT. 2010. A human-specific deletion in mouse Cmah increases disease severity in the mdx model of Duchenne muscular dystrophy. Sci Transl Med. 2:42-54). Here, we demonstrate similar changes in cardiac and skeletal muscle pathology and physiology resulting from Cmah deletion in α-sarcoglycan-deficient (Sgca(-/-)) mice, a model for limb girdle muscular dystrophy 2D. These experiments demonstrate that loss of mouse Cmah can worsen disease severity in more than one form of muscular dystrophy and suggest that Cmah may be a general genetic modifier of muscle disease.
Assuntos
Músculo Esquelético/patologia , Miocárdio/patologia , Ácidos Neuramínicos/metabolismo , Sarcoglicanas/genética , Animais , Deleção de Genes , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiopatologia , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismo , Miocárdio/metabolismo , Sarcoglicanas/deficiênciaRESUMO
BACKGROUND: Prostate tumors shed circulating tumor cells (CTCs) into the blood stream. Increased evidence shows that CTCs are often present in metastatic prostate cancer and can be alternative sources for disease profiling and prognostication. Here we postulate that CTCs expressing genes related to epithelial-mesenchymal transition (EMT) are strong predictors of metastatic prostate cancer. METHODS: A microfiltration system was used to trap CTCs from peripheral blood based on size selection of large epithelial-like cells without CD45 leukocyte marker. These cells individually retrieved with a micromanipulator device were assessed for cell membrane physical properties using atomic force microscopy. Additionally, 38 CTCs from eight prostate cancer patients were used to determine expression profiles of 84 EMT-related and reference genes using a microfluidics-based PCR system. RESULTS: Increased cell elasticity and membrane smoothness were found in CTCs compared to noncancerous cells, highlighting their potential invasiveness and mobility in the peripheral circulation. Despite heterogeneous expression patterns of individual CTCs, genes that promote mesenchymal transitioning into a more malignant state, including IGF1, IGF2, EGFR, FOXP3, and TGFB3, were commonly observed in these cells. An additional subset of EMT-related genes (e.g., PTPRN2, ALDH1, ESR2, and WNT5A) were expressed in CTCs of castration-resistant cancer, but less frequently in castration-sensitive cancer. CONCLUSIONS: The study suggests that an incremental expression of EMT-related genes in CTCs is associated with metastatic castration-resistant cancer. Although CTCs represent a group of highly heterogeneous cells, their unique EMT-related gene signatures provide a new opportunity for personalized treatments with targeted inhibitors in advanced prostate cancer patients.
Assuntos
Transição Epitelial-Mesenquimal/genética , Neoplasias Hormônio-Dependentes/metabolismo , Células Neoplásicas Circulantes/metabolismo , Neoplasias da Próstata/metabolismo , Linhagem Celular Tumoral , DNA de Neoplasias/química , DNA de Neoplasias/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Masculino , Técnicas Analíticas Microfluídicas , Microscopia de Força Atômica , Neoplasias Hormônio-Dependentes/sangue , Neoplasias Hormônio-Dependentes/genética , Células Neoplásicas Circulantes/patologia , Neoplasias da Próstata/sangue , Neoplasias da Próstata/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Célula Única/métodosRESUMO
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.
Assuntos
Antígeno CD47 , Macrófagos , Metástase Neoplásica , Fagocitose , Proteínas Proto-Oncogênicas c-myc , Evasão Tumoral , Humanos , Masculino , Proteínas de Transporte , Antígeno CD47/metabolismo , Macrófagos/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/imunologia , Transdução de Sinais , Evasão Tumoral/genética , Evasão Tumoral/imunologia , Metástase Neoplásica/genética , Metástase Neoplásica/imunologia , Células Tumorais CultivadasRESUMO
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.
Assuntos
Neoplasias Pulmonares , Macrófagos Associados a Tumor , Humanos , Animais , Camundongos , Células Endoteliais , Transdução de Sinais , Diferenciação Celular , Microambiente Tumoral , Linhagem Celular TumoralRESUMO
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.
Assuntos
Cromatina/metabolismo , Glutaratos/metabolismo , Oxirredutases do Álcool/metabolismo , Ácido Ascórbico/análogos & derivados , Ácido Ascórbico/metabolismo , Diferenciação Celular , Linhagem Celular Tumoral , Reparo do DNA/fisiologia , Epigenoma/genética , Fatores de Transcrição Forkhead/genética , Expressão Gênica/genética , Regulação da Expressão Gênica/genética , Humanos , Isocitrato Desidrogenase/genética , Neoplasias/genética , Neoplasias/metabolismo , Nucleossomos/metabolismo , Proteínas Repressoras/genéticaRESUMO
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.
Assuntos
Biomarcadores/metabolismo , Endometriose/metabolismo , Endométrio/citologia , Epitélio/metabolismo , Junções Intercelulares/metabolismo , Proteômica/métodos , Células Cultivadas , Técnicas de Cocultura , Biologia Computacional , Endométrio/metabolismo , Endométrio/patologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Humanos , Análise de Célula Única , Células Estromais/citologia , Células Estromais/metabolismoRESUMO
Vaccination has become an important therapeutic approach to the treatment of Alzheimer's disease (AD), however, immunization with Abeta amyloid can have unwanted, potentially lethal, side effects. Here we demonstrate an alternative peptide-mimotope vaccine strategy using the SDPM1 peptide. SDPM1 is a 20 amino acid peptide bounded by cysteines that binds tetramer forms of Abeta(1-40)- and Abeta(1-42)-amyloids and blocks subsequent Abeta amyloid aggregation. Immunization of mice with SDPM1 induced peptide-mimotope antibodies with the same biological activity as the SDPM1 peptide. When done prior to the onset of amyloid plaque formation, SDPM1 vaccination of APPswePSEN1(A246E) transgenic mice reduced amyloid plaque burden and Abeta(1-40) and Abeta(1-42) levels in the brain, improved cognitive performance in Morris water maze tests, and resulted in no increased T cell responses to immunogenic or Abeta peptides or brain inflammation. When done after plaque burden was already significant, SDPM1 immunization still significantly reduced amyloid plaque burden and Abeta(1-40/1-42) peptide levels in APPswePSEN1(A246E) brain without inducing encephalitogenic T cell responses or brain inflammation, but treatment at this stage did not improve cognitive function. These experiments demonstrate the efficacy of a novel vaccine approach for Alzheimer's disease where immunization with an Abeta(1-40/1-42) amyloid-specific binding and blocking peptide is used to inhibit the development of neuropathology and cognitive dysfunction.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Vacinas contra Alzheimer/uso terapêutico , Encéfalo/imunologia , Cognição/efeitos dos fármacos , Aprendizagem em Labirinto/efeitos dos fármacos , Placa Amiloide/imunologia , Doença de Alzheimer/imunologia , Doença de Alzheimer/patologia , Doença de Alzheimer/fisiopatologia , Vacinas contra Alzheimer/imunologia , Peptídeos beta-Amiloides , Análise de Variância , Animais , Western Blotting , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Encéfalo/fisiopatologia , Modelos Animais de Doenças , Imuno-Histoquímica , Imunoprecipitação , Camundongos , Camundongos Transgênicos , Fragmentos de Peptídeos , Placa Amiloide/efeitos dos fármacos , Placa Amiloide/patologia , VacinaçãoRESUMO
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.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/genética , Cromossomos Humanos Par 17/genética , Cromotripsia , Receptor alfa de Estrogênio/metabolismo , Regulação Neoplásica da Expressão Gênica , Biomarcadores Tumorais/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ciclo Celular , Proliferação de Células , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Prognóstico , Taxa de Sobrevida , Transcrição Gênica , Células Tumorais Cultivadas , Sequenciamento do Exoma , Sequenciamento Completo do GenomaRESUMO
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.
Assuntos
Neoplasias do Endométrio/metabolismo , Moléculas de Adesão Juncional/metabolismo , Obesidade/metabolismo , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Proteína Smad4/metabolismo , Tecido Adiposo/patologia , Regulação para Baixo/genética , Neoplasias do Endométrio/complicações , Neoplasias do Endométrio/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Proteína-1 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Obesidade/complicações , Ligação Proteica , Proteólise , Proteômica , Proteína Smad4/genética , Células Estromais/metabolismo , Transcrição Gênica , Fator de Crescimento Transformador beta/metabolismo , Microambiente Tumoral , Ubiquitina/metabolismoRESUMO
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.
Assuntos
Membro C3 da Família 1 de alfa-Ceto Redutase/metabolismo , Carcinoma/enzimologia , Neoplasias da Próstata/enzimologia , Sulfotransferases/metabolismo , Animais , Transição Epitelial-Mesenquimal , Humanos , Masculino , Camundongos Nus , Metástase Neoplásica , Transplante de Neoplasias , Receptores Androgênicos/metabolismoRESUMO
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.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Janus Quinase 1/antagonistas & inibidores , Neoplasias Pulmonares/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Idoso , Idoso de 80 Anos ou mais , Animais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Sinergismo Farmacológico , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Estudos de Viabilidade , Feminino , Citometria de Fluxo/métodos , Humanos , Janus Quinase 1/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Nitrilas , Inibidores de Proteínas Quinases/uso terapêutico , Proteômica/métodos , Proteínas Proto-Oncogênicas/metabolismo , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , RNA-Seq , Receptores Proteína Tirosina Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Análise de Célula Única/métodos , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Análise Serial de Tecidos , Ensaios Antitumorais Modelo de Xenoenxerto , Receptor Tirosina Quinase AxlRESUMO
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.
Assuntos
Carcinogênese/genética , Metilação de DNA , Neoplasias Hepáticas/genética , Sequências Reguladoras de Ácido Nucleico/genética , Animais , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Carcinoma Hepatocelular/genética , Proteínas de Ciclo Celular , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Desmetilação , Epigênese Genética , Deleção de Genes , Regulação Neoplásica da Expressão Gênica , Humanos , Fígado , Camundongos , Camundongos Transgênicos , Proteínas Nucleares/metabolismo , Prognóstico , Regiões Promotoras Genéticas , Transativadores , Fatores de Transcrição/metabolismo , Ativação Transcricional , Regulação para Cima , Proteínas Virais Reguladoras e AcessóriasRESUMO
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.