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1.
Cell ; 186(13): 2765-2782.e28, 2023 06 22.
Artículo en Inglés | MEDLINE | ID: mdl-37327786

RESUMEN

Cancer is characterized by hypomethylation-associated silencing of large chromatin domains, whose contribution to tumorigenesis is uncertain. Through high-resolution genome-wide single-cell DNA methylation sequencing, we identify 40 core domains that are uniformly hypomethylated from the earliest detectable stages of prostate malignancy through metastatic circulating tumor cells (CTCs). Nested among these repressive domains are smaller loci with preserved methylation that escape silencing and are enriched for cell proliferation genes. Transcriptionally silenced genes within the core hypomethylated domains are enriched for immune-related genes; prominent among these is a single gene cluster harboring all five CD1 genes that present lipid antigens to NKT cells and four IFI16-related interferon-inducible genes implicated in innate immunity. The re-expression of CD1 or IFI16 murine orthologs in immuno-competent mice abrogates tumorigenesis, accompanied by the activation of anti-tumor immunity. Thus, early epigenetic changes may shape tumorigenesis, targeting co-located genes within defined chromosomal loci. Hypomethylation domains are detectable in blood specimens enriched for CTCs.


Asunto(s)
Metilación de ADN , Neoplasias de la Próstata , Animales , Humanos , Masculino , Ratones , Carcinogénesis/genética , ADN , Epigénesis Genética , Neoplasias de la Próstata/genética , Células Neoplásicas Circulantes
2.
Cell ; 158(5): 1110-1122, 2014 Aug 28.
Artículo en Inglés | MEDLINE | ID: mdl-25171411

RESUMEN

Circulating tumor cell clusters (CTC clusters) are present in the blood of patients with cancer but their contribution to metastasis is not well defined. Using mouse models with tagged mammary tumors, we demonstrate that CTC clusters arise from oligoclonal tumor cell groupings and not from intravascular aggregation events. Although rare in the circulation compared with single CTCs, CTC clusters have 23- to 50-fold increased metastatic potential. In patients with breast cancer, single-cell resolution RNA sequencing of CTC clusters and single CTCs, matched within individual blood samples, identifies the cell junction component plakoglobin as highly differentially expressed. In mouse models, knockdown of plakoglobin abrogates CTC cluster formation and suppresses lung metastases. In breast cancer patients, both abundance of CTC clusters and high tumor plakoglobin levels denote adverse outcomes. Thus, CTC clusters are derived from multicellular groupings of primary tumor cells held together through plakoglobin-dependent intercellular adhesion, and though rare, they greatly contribute to the metastatic spread of cancer.


Asunto(s)
Neoplasias de la Mama/patología , Metástasis de la Neoplasia/patología , Células Neoplásicas Circulantes/patología , Animales , Neoplasias de la Mama/fisiopatología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Neoplasias de la Próstata/patología , Neoplasias de la Próstata/fisiopatología , Análisis de Secuencia de ARN , Análisis de la Célula Individual , gamma Catenina/metabolismo
3.
Mol Cell ; 81(19): 4041-4058.e15, 2021 10 07.
Artículo en Inglés | MEDLINE | ID: mdl-34624217

RESUMEN

Deregulation of oncogenic signals in cancer triggers replication stress. Immediate early genes (IEGs) are rapidly and transiently expressed following stressful signals, contributing to an integrated response. Here, we find that the orphan nuclear receptor NR4A1 localizes across the gene body and 3' UTR of IEGs, where it inhibits transcriptional elongation by RNA Pol II, generating R-loops and accessible chromatin domains. Acute replication stress causes immediate dissociation of NR4A1 and a burst of transcriptionally poised IEG expression. Ectopic expression of NR4A1 enhances tumorigenesis by breast cancer cells, while its deletion leads to massive chromosomal instability and proliferative failure, driven by deregulated expression of its IEG target, FOS. Approximately half of breast and other primary cancers exhibit accessible chromatin domains at IEG gene bodies, consistent with this stress-regulatory pathway. Cancers that have retained this mechanism in adapting to oncogenic replication stress may be dependent on NR4A1 for their proliferation.


Asunto(s)
Neoplasias de la Mama/metabolismo , Proliferación Celular , Proteínas Inmediatas-Precoces/metabolismo , Mitosis , Células Neoplásicas Circulantes/metabolismo , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , Regiones no Traducidas 3' , Animales , Antineoplásicos/farmacología , Sitios de Unión , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Proliferación Celular/efectos de los fármacos , Ensamble y Desensamble de Cromatina , Femenino , Regulación Neoplásica de la Expresión Génica , Inestabilidad Genómica , Células HEK293 , Humanos , Proteínas Inmediatas-Precoces/genética , Indoles/farmacología , Células MCF-7 , Ratones Endogámicos NOD , Ratones SCID , Mitosis/efectos de los fármacos , Células Neoplásicas Circulantes/efectos de los fármacos , Células Neoplásicas Circulantes/patología , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares/antagonistas & inhibidores , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Fenilacetatos/farmacología , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , Estructuras R-Loop , ARN Polimerasa II/genética , ARN Polimerasa II/metabolismo , Transducción de Señal , Elongación de la Transcripción Genética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto
4.
Cell ; 141(1): 69-80, 2010 Apr 02.
Artículo en Inglés | MEDLINE | ID: mdl-20371346

RESUMEN

Accumulating evidence implicates heterogeneity within cancer cell populations in the response to stressful exposures, including drug treatments. While modeling the acute response to various anticancer agents in drug-sensitive human tumor cell lines, we consistently detected a small subpopulation of reversibly "drug-tolerant" cells. These cells demonstrate >100-fold reduced drug sensitivity and maintain viability via engagement of IGF-1 receptor signaling and an altered chromatin state that requires the histone demethylase RBP2/KDM5A/Jarid1A. This drug-tolerant phenotype is transiently acquired and relinquished at low frequency by individual cells within the population, implicating the dynamic regulation of phenotypic heterogeneity in drug tolerance. The drug-tolerant subpopulation can be selectively ablated by treatment with IGF-1 receptor inhibitors or chromatin-modifying agents, potentially yielding a therapeutic opportunity. Together, these findings suggest that cancer cell populations employ a dynamic survival strategy in which individual cells transiently assume a reversibly drug-tolerant state to protect the population from eradication by potentially lethal exposures.


Asunto(s)
Resistencia a Antineoplásicos , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Línea Celular Tumoral , Cromatina/metabolismo , Cromatina/patología , Daño del ADN , Inhibidores de Histona Desacetilasas/farmacología , Histona Demetilasas/metabolismo , Humanos , Histona Demetilasas con Dominio de Jumonji/antagonistas & inhibidores , Histona Demetilasas con Dominio de Jumonji/genética , Histona Demetilasas con Dominio de Jumonji/metabolismo , Neoplasias/metabolismo , Receptor IGF Tipo 1/metabolismo
5.
Breast Cancer Res Treat ; 201(1): 43-56, 2023 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-37318638

RESUMEN

PURPOSE: Metastatic hormone receptor-positive (HR+) breast cancer initially responds to serial courses of endocrine therapy, but ultimately becomes refractory. Elacestrant, a new generation FDA-approved oral selective estrogen receptor degrader (SERD) and antagonist, has demonstrated efficacy in a subset of women with advanced HR+breast cancer, but there are few patient-derived models to characterize its effect in advanced cancers with diverse treatment histories and acquired mutations. METHODS: We analyzed clinical outcomes with elacestrant, compared with endocrine therapy, among women who had previously been treated with a fulvestrant-containing regimen from the recent phase 3 EMERALD Study. We further modeled sensitivity to elacestrant, compared with the currently approved SERD, fulvestrant in patient-derived xenograft (PDX) models and cultured circulating tumor cells (CTCs). RESULTS: Analysis of the subset of breast cancer patients enrolled in the EMERALD study who had previously received a fulvestrant-containing regimen indicates that they had better progression-free survival with elacestrant than with standard-of-care endocrine therapy, a finding that was independent estrogen receptor (ESR1) gene mutations. We modeled elacestrant responsiveness using patient-derived xenograft (PDX) models and in ex vivo cultured CTCs derived from patients with HR+breast cancer extensively treated with multiple endocrine therapies, including fulvestrant. Both CTCs and PDX models are refractory to fulvestrant but sensitive to elacestrant, independent of mutations in ESR1 and Phosphatidylinositol-4,5-Bisphosphate 3-Kinase Catalytic Subunit Alpha (PIK3CA) genes. CONCLUSION: Elacestrant retains efficacy in breast cancer cells that have acquired resistance to currently available ER targeting therapies. Elacestrant may be an option for patients with HR+/HER2- breast cancer whose disease progressed on fulvestrant in the metastatic setting. TRANSLATIONAL RELEVANCE: Serial endocrine therapy is the mainstay of management for metastatic HR+breast cancer, but acquisition of drug resistance highlights the need for better therapies. Elacestrant is a recently FDA-approved novel oral selective estrogen receptor degrader (SERD), with demonstrated efficacy in the EMERALD phase 3 clinical trial of refractory HR+breast cancer. Subgroup analysis of the EMERALD clinical trial identifies clinical benefit with elacestrant in patients who had received prior fulvestrant independent of the mutational status of the ESR1 gene, supporting its potential utility in treating refractory HR+breast cancer. Here, we use pre-clinical models, including ex vivo cultures of circulating tumor cells and patient-derived xenografts, to demonstrate the efficacy of elacestrant in breast cancer cells with acquired resistance to fulvestrant.


Asunto(s)
Neoplasias de la Mama , Células Neoplásicas Circulantes , Animales , Humanos , Femenino , Fulvestrant , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Receptores de Estrógenos , Antagonistas de Estrógenos/uso terapéutico , Modelos Animales de Enfermedad , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico
6.
Nature ; 537(7618): 102-106, 2016 09 01.
Artículo en Inglés | MEDLINE | ID: mdl-27556950

RESUMEN

Circulating tumour cells in women with advanced oestrogen-receptor (ER)-positive/human epidermal growth factor receptor 2 (HER2)-negative breast cancer acquire a HER2-positive subpopulation after multiple courses of therapy. In contrast to HER2-amplified primary breast cancer, which is highly sensitive to HER2-targeted therapy, the clinical significance of acquired HER2 heterogeneity during the evolution of metastatic breast cancer is unknown. Here we analyse circulating tumour cells from 19 women with ER+/HER2- primary tumours, 84% of whom had acquired circulating tumour cells expressing HER2. Cultured circulating tumour cells maintain discrete HER2+ and HER2- subpopulations: HER2+ circulating tumour cells are more proliferative but not addicted to HER2, consistent with activation of multiple signalling pathways; HER2- circulating tumour cells show activation of Notch and DNA damage pathways, exhibiting resistance to cytotoxic chemotherapy, but sensitivity to Notch inhibition. HER2+ and HER2- circulating tumour cells interconvert spontaneously, with cells of one phenotype producing daughters of the opposite within four cell doublings. Although HER2+ and HER2- circulating tumour cells have comparable tumour initiating potential, differential proliferation favours the HER2+ state, while oxidative stress or cytotoxic chemotherapy enhances transition to the HER2- phenotype. Simultaneous treatment with paclitaxel and Notch inhibitors achieves sustained suppression of tumorigenesis in orthotopic circulating tumour cell-derived tumour models. Together, these results point to distinct yet interconverting phenotypes within patient-derived circulating tumour cells, contributing to progression of breast cancer and acquisition of drug resistance.


Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Células Neoplásicas Circulantes/metabolismo , Células Neoplásicas Circulantes/patología , Receptor ErbB-2/metabolismo , Neoplasias de la Mama/tratamiento farmacológico , Línea Celular Tumoral , Proliferación Celular , Resistencia a Antineoplásicos , Femenino , Humanos , Células Neoplásicas Circulantes/efectos de los fármacos , Fenotipo , Receptor ErbB-2/deficiencia , Receptor Notch1/antagonistas & inhibidores , Receptor Notch1/metabolismo , Transducción de Señal
7.
Proc Natl Acad Sci U S A ; 116(12): 5223-5232, 2019 03 19.
Artículo en Inglés | MEDLINE | ID: mdl-30819896

RESUMEN

Tumor-stromal communication within the microenvironment contributes to initiation of metastasis and may present a therapeutic opportunity. Using serial single-cell RNA sequencing in an orthotopic mouse prostate cancer model, we find up-regulation of prolactin receptor as cancer cells that have disseminated to the lungs expand into micrometastases. Secretion of the ligand prolactin by adjacent lung stromal cells is induced by tumor cell production of the COX-2 synthetic product prostaglandin E2 (PGE2). PGE2 treatment of fibroblasts activates the orphan nuclear receptor NR4A (Nur77), with prolactin as a major transcriptional target for the NR4A-retinoid X receptor (RXR) heterodimer. Ectopic expression of prolactin receptor in mouse cancer cells enhances micrometastasis, while treatment with the COX-2 inhibitor celecoxib abrogates prolactin secretion by fibroblasts and reduces tumor initiation. Across multiple human cancers, COX-2, prolactin, and prolactin receptor show consistent differential expression in tumor and stromal compartments. Such paracrine cross-talk may thus contribute to the documented efficacy of COX-2 inhibitors in cancer suppression.


Asunto(s)
Carcinogénesis/metabolismo , Prolactina/metabolismo , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Transducción de Señal/fisiología , Células del Estroma/metabolismo , Animales , Carcinogénesis/efectos de los fármacos , Celecoxib/farmacología , Transformación Celular Neoplásica/efectos de los fármacos , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Ciclooxigenasa 2/metabolismo , Inhibidores de la Ciclooxigenasa 2/farmacología , Dinoprostona/metabolismo , Modelos Animales de Enfermedad , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismo , Humanos , Masculino , Ratones , Miembro 1 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Receptores X Retinoide/metabolismo , Transducción de Señal/efectos de los fármacos , Células del Estroma/efectos de los fármacos , Células del Estroma/patología , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiología
8.
Breast Cancer Res Treat ; 188(1): 43-52, 2021 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-34101078

RESUMEN

PURPOSE: Therapeutic efficacy of hormonal therapies to target estrogen receptor (ER)-positive breast cancer is limited by the acquisition of ligand-independent ESR1 mutations, which confer treatment resistance to aromatase inhibitors (AIs). Monitoring for the emergence of such mutations may enable individualized therapy. We thus assessed CTC- and ctDNA-based detection of ESR1 mutations with the aim of evaluating non-invasive approaches for the determination of endocrine resistance. PATIENTS AND METHODS: In a prospective cohort of 55 women with hormone receptor-positive metastatic breast cancer, we isolated circulating tumor cells (CTCs) and developed a high-sensitivity method for the detection of ESR1 mutations in these CTCs. In patients with sufficient plasma for the simultaneous extraction of circulating tumor DNA (ctDNA), we performed a parallel analysis of ESR1 mutations using multiplex droplet digital PCR (ddPCR) and examined the agreement between these two platforms. Finally, we isolated single CTCs from a subset of these patients and reviewed RNA expression to explore alternate methods of evaluating endocrine responsiveness. RESULTS: High-sensitivity ESR1 sequencing from CTCs revealed mono- and oligoclonal mutations in 22% of patients. These were concordant with plasma DNA sequencing in 95% of cases. Emergence of ESR1 mutations was correlated both with time to metastatic relapse and duration of AI therapy following such recurrence. The Presence of an ESR1 mutation, compared to ESR1 wild type, was associated with markedly shorter Progression-Free Survival on AI-based therapies (p = 0.0006), but unaltered to other non-AI-based therapies (p = 0.73). Compared with ESR1 mutant cases, AI-resistant CTCs with wild-type ESR1 showed an elevated ER-coactivator RNA signature, consistent with their predicted response to second-line hormonal therapies. CONCLUSION: Blood-based serial monitoring may guide the selection of precision therapeutics for women with AI-resistant ER-positive breast cancer.


Asunto(s)
Neoplasias de la Mama , ADN Tumoral Circulante , Células Neoplásicas Circulantes , Receptor alfa de Estrógeno/genética , Femenino , Genotipo , Humanos , Mutación , Recurrencia Local de Neoplasia , Estudios Prospectivos
9.
Proc Natl Acad Sci U S A ; 115(10): 2467-2472, 2018 03 06.
Artículo en Inglés | MEDLINE | ID: mdl-29453278

RESUMEN

A subset of patients with metastatic melanoma have sustained remissions following treatment with immune checkpoint inhibitors. However, analyses of pretreatment tumor biopsies for markers predictive of response, including PD-1 ligand (PD-L1) expression and mutational burden, are insufficiently precise to guide treatment selection, and clinical radiographic evidence of response on therapy may be delayed, leading to some patients receiving potentially ineffective but toxic therapy. Here, we developed a molecular signature of melanoma circulating tumor cells (CTCs) to quantify early tumor response using blood-based monitoring. A quantitative 19-gene digital RNA signature (CTC score) applied to microfluidically enriched CTCs robustly distinguishes melanoma cells, within a background of blood cells in reconstituted and in patient-derived (n = 42) blood specimens. In a prospective cohort of 49 patients treated with immune checkpoint inhibitors, a decrease in CTC score within 7 weeks of therapy correlates with marked improvement in progression-free survival [hazard ratio (HR), 0.17; P = 0.008] and overall survival (HR, 0.12; P = 0.04). Thus, digital quantitation of melanoma CTC-derived transcripts enables serial noninvasive monitoring of tumor burden, supporting the rational application of immune checkpoint inhibition therapies.


Asunto(s)
Antineoplásicos Inmunológicos , Biomarcadores de Tumor/sangre , Melanoma , Células Neoplásicas Circulantes , Neoplasias Cutáneas , Antineoplásicos Inmunológicos/farmacología , Antineoplásicos Inmunológicos/uso terapéutico , Biomarcadores de Tumor/química , Tratamiento Basado en Trasplante de Células y Tejidos , Femenino , Humanos , Estimación de Kaplan-Meier , Biopsia Líquida , Masculino , Melanoma/sangre , Melanoma/diagnóstico , Melanoma/tratamiento farmacológico , Melanoma/mortalidad , Persona de Mediana Edad , Células Neoplásicas Circulantes/química , Células Neoplásicas Circulantes/efectos de los fármacos , ARN/análisis , ARN/genética , ARN/metabolismo , Neoplasias Cutáneas/sangre , Neoplasias Cutáneas/diagnóstico , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/mortalidad
10.
Nat Methods ; 12(7): 685-91, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-25984697

RESUMEN

Cancer cells metastasize through the bloodstream either as single migratory circulating tumor cells (CTCs) or as multicellular groupings (CTC clusters). Existing technologies for CTC enrichment are designed to isolate single CTCs, and although CTC clusters are detectable in some cases, their true prevalence and significance remain to be determined. Here we developed a microchip technology (the Cluster-Chip) to capture CTC clusters independently of tumor-specific markers from unprocessed blood. CTC clusters are isolated through specialized bifurcating traps under low-shear stress conditions that preserve their integrity, and even two-cell clusters are captured efficiently. Using the Cluster-Chip, we identified CTC clusters in 30-40% of patients with metastatic breast or prostate cancer or with melanoma. RNA sequencing of CTC clusters confirmed their tumor origin and identified tissue-derived macrophages within the clusters. Efficient capture of CTC clusters will enable the detailed characterization of their biological properties and role in metastasis.


Asunto(s)
Técnicas Analíticas Microfluídicas , Células Neoplásicas Circulantes , Neoplasias de la Mama/patología , Línea Celular Tumoral , Femenino , Humanos , Inmunohistoquímica , Masculino , Neoplasias de la Próstata/patología , Análisis de Secuencia de ARN
11.
Nature ; 487(7408): 510-3, 2012 Jul 26.
Artículo en Inglés | MEDLINE | ID: mdl-22763454

RESUMEN

Circulating tumour cells (CTCs) shed into blood from primary cancers include putative precursors that initiate distal metastases. Although these cells are extraordinarily rare, they may identify cellular pathways contributing to the blood-borne dissemination of cancer. Here, we adapted a microfluidic device for efficient capture of CTCs from an endogenous mouse pancreatic cancer model and subjected CTCs to single-molecule RNA sequencing, identifying Wnt2 as a candidate gene enriched in CTCs. Expression of WNT2 in pancreatic cancer cells suppresses anoikis, enhances anchorage-independent sphere formation, and increases metastatic propensity in vivo. This effect is correlated with fibronectin upregulation and suppressed by inhibition of MAP3K7 (also known as TAK1) kinase. In humans, formation of non-adherent tumour spheres by pancreatic cancer cells is associated with upregulation of multiple WNT genes, and pancreatic CTCs revealed enrichment for WNT signalling in 5 out of 11 cases. Thus, molecular analysis of CTCs may identify candidate therapeutic targets to prevent the distal spread of cancer.


Asunto(s)
Regulación Neoplásica de la Expresión Génica/genética , Metástasis de la Neoplasia/genética , Células Neoplásicas Circulantes/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patología , Proteínas Wnt/metabolismo , Vía de Señalización Wnt/genética , Animales , Supervivencia Celular , Inhibición de Contacto , Modelos Animales de Enfermedad , Genes Relacionados con las Neoplasias/genética , Humanos , Quinasas Quinasa Quinasa PAM/antagonistas & inhibidores , Ratones , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Análisis de Secuencia de ARN , Proteínas Wnt/genética , Proteína wnt2/genética , Proteína wnt2/metabolismo
12.
Proc Natl Acad Sci U S A ; 112(49): 15148-53, 2015 Dec 08.
Artículo en Inglés | MEDLINE | ID: mdl-26575630

RESUMEN

Aberrant transcription of the pericentromeric human satellite II (HSATII) repeat is present in a wide variety of epithelial cancers. In deriving experimental systems to study its deregulation, we observed that HSATII expression is induced in colon cancer cells cultured as xenografts or under nonadherent conditions in vitro, but it is rapidly lost in standard 2D cultures. Unexpectedly, physiological induction of endogenous HSATII RNA, as well as introduction of synthetic HSATII transcripts, generated cDNA intermediates in the form of DNA/RNA hybrids. Single molecule sequencing of tumor xenografts showed that HSATII RNA-derived DNA (rdDNA) molecules are stably incorporated within pericentromeric loci. Suppression of RT activity using small molecule inhibitors reduced HSATII copy gain. Analysis of whole-genome sequencing data revealed that HSATII copy number gain is a common feature in primary human colon tumors and is associated with a lower overall survival. Together, our observations suggest that cancer-associated derepression of specific repetitive sequences can promote their RNA-driven genomic expansion, with potential implications on pericentromeric architecture.


Asunto(s)
Centrómero/genética , ADN Satélite/genética , Neoplasias/genética , Secuencias Repetitivas de Ácidos Nucleicos , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Humanos , Hibridación de Ácido Nucleico , ARN/genética
13.
Genes Dev ; 23(15): 1737-42, 2009 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-19651985

RESUMEN

Epithelial-to-mesenchymal transition (EMT) plays an important role during normal embryogenesis, and it has been implicated in cancer invasion and metastasis. Here, we report that Ladybird homeobox 1 (LBX1), a developmentally regulated homeobox gene, directs expression of the known EMT inducers ZEB1, ZEB2, Snail1, and transforming growth factor beta2 (TGFB2). In mammary epithelial cells, overexpression of LBX1 leads to morphological transformation, expression of mesenchymal markers, enhanced cell migration, increased CD44(high)/CD24(low) progenitor cell population, and tumorigenic cooperation with known oncogenes. In human breast cancer, LBX1 is up-regulated in the unfavorable estrogen receptor (ER)/progesterone (PR)/HER2 triple-negative basal-like subtype. Thus, aberrant expression of LBX1 may lead to the activation of a developmentally regulated EMT pathway in human breast cancer.


Asunto(s)
Neoplasias de la Mama/fisiopatología , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica , Proteínas de Homeodominio/metabolismo , Proteínas de Transporte de Catión Orgánico/metabolismo , Factores de Transcripción/metabolismo , Animales , Antígeno CD24/metabolismo , Línea Celular Tumoral , Movimiento Celular , Proteínas de Homeodominio/genética , Humanos , Receptores de Hialuranos/metabolismo , Glándulas Mamarias Humanas/citología , Glándulas Mamarias Humanas/metabolismo , Oncogenes/fisiología , Proteínas Represoras/metabolismo , Factores de Transcripción de la Familia Snail , Células Madre/metabolismo , Factores de Transcripción/genética , Factor de Crecimiento Transformador beta2/metabolismo , Regulación hacia Arriba , Caja Homeótica 2 de Unión a E-Box con Dedos de Zinc , Homeobox 1 de Unión a la E-Box con Dedos de Zinc
14.
J Biol Chem ; 290(23): 14381-90, 2015 Jun 05.
Artículo en Inglés | MEDLINE | ID: mdl-25882849

RESUMEN

WTX encodes a tumor suppressor implicated in the pediatric kidney cancer Wilms tumor and in mesenchymal differentiation with potentially distinct functions in the cytoplasm, at the plasma membrane, and in the nucleus. Although modulating components of the WNT signaling pathway is a proposed function for cytoplasmic and membrane-bound WTX, its nuclear properties are not well understood. Here we report that the transcriptional corepressor TRIM28 is the major binding partner for nuclear WTX. WTX interacted with the coiled coil domain of TRIM28 required for its binding to Krüppel-associated box domains of transcription factors and for its chromatin recruitment through its own coiled coil and proline-rich domains. Knockdown of endogenous WTX reduced the recruitment of TRIM28 to a chromatinized reporter sequence and its ability to repress a target transcript. In mouse embryonic stem cells where TRIM28 plays a major role in repressing endogenous retroviruses and long interspersed elements, knockdown of either TRIM28 or WTX combined with single molecule RNA sequencing revealed a highly significant shared set of differentially regulated transcripts, including derepression of non-coding repetitive sequences and their neighboring protein encoding genes (p < 1e-20). In mesenchymal precursor cells, depletion of WTX and TRIM28 resulted in analogous ß-catenin-independent defects in adipogenic and osteogenic differentiation, and knockdown of WTX reduced TRIM28 binding to Pparγ promoter. Together, the physical and functional interaction between WTX and TRIM28 suggests that the nuclear fraction of WTX plays a role in epigenetic silencing, an effect that may contribute to its function as a regulator of cellular differentiation and tumorigenesis.


Asunto(s)
Células Madre Embrionarias/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Represoras/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Adipogénesis , Animales , Línea Celular , Cromatina/genética , Cromatina/metabolismo , Células Madre Embrionarias/citología , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Ratones , Proteínas Nucleares/genética , Mapas de Interacción de Proteínas , Proteínas Represoras/genética , Activación Transcripcional , Proteína 28 que Contiene Motivos Tripartito , Proteínas Supresoras de Tumor/genética , Tumor de Wilms/genética , Tumor de Wilms/metabolismo
15.
Proc Natl Acad Sci U S A ; 110(13): 5139-44, 2013 Mar 26.
Artículo en Inglés | MEDLINE | ID: mdl-23483055

RESUMEN

Metastasis confronts clinicians with two major challenges: estimating the patient's risk of metastasis and identifying therapeutic targets. Because they are key signal integrators connecting cellular processes to clinical outcome, we aimed to identify transcriptional nodes regulating cancer cell metastasis. Using rodent xenograft models that we previously developed, we identified the transcription factor Fos-related antigen-1 (Fra-1) as a key coordinator of metastasis. Because Fra-1 often is overexpressed in human metastatic breast cancers and has been shown to control their invasive potential in vitro, we aimed to assess the implication and prognostic significance of the Fra-1-dependent genetic program in breast cancer metastasis and to identify potential Fra-1-dependent therapeutic targets. In several in vivo assays in mice, we demonstrate that stable RNAi depletion of Fra-1 from human breast cancer cells strongly suppresses their ability to metastasize. These results support a clinically important role for Fra-1 and the genetic program it controls. We show that a Fra-1-dependent gene-expression signature accurately predicts recurrence of breast cancer. Furthermore, a synthetic lethal drug screen revealed that antagonists of the adenosine receptor A2B (ADORA2B) are preferentially toxic to breast tumor cells expressing Fra-1. Both RNAi silencing and pharmacologic blockade of ADORA2B inhibited filopodia formation and invasive activity of breast cancer cells and correspondingly reduced tumor outgrowth in the lungs. These data show that Fra-1 activity is causally involved in and is a prognostic indicator of breast cancer metastasis. They suggest that Fra-1 activity predicts responsiveness to inhibition of pharmacologically tractable targets, such as ADORA2B, which may be used for clinical interference of metastatic breast cancer.


Asunto(s)
Neoplasias de la Mama/metabolismo , Regulación Neoplásica de la Expresión Génica , Proteínas Proto-Oncogénicas c-fos/metabolismo , Receptor de Adenosina A2B/metabolismo , Antagonistas del Receptor de Adenosina A2/farmacología , Animales , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Invasividad Neoplásica , Metástasis de la Neoplasia , Trasplante de Neoplasias , Proteínas Proto-Oncogénicas c-fos/genética , Seudópodos/genética , Seudópodos/metabolismo , Seudópodos/patología , Ratas , Receptor de Adenosina A2B/genética , Trasplante Heterólogo , Ensayos Antitumor por Modelo de Xenoinjerto
16.
Nature ; 462(7269): 108-12, 2009 Nov 05.
Artículo en Inglés | MEDLINE | ID: mdl-19847166

RESUMEN

The proto-oncogene KRAS is mutated in a wide array of human cancers, most of which are aggressive and respond poorly to standard therapies. Although the identification of specific oncogenes has led to the development of clinically effective, molecularly targeted therapies in some cases, KRAS has remained refractory to this approach. A complementary strategy for targeting KRAS is to identify gene products that, when inhibited, result in cell death only in the presence of an oncogenic allele. Here we have used systematic RNA interference to detect synthetic lethal partners of oncogenic KRAS and found that the non-canonical IkappaB kinase TBK1 was selectively essential in cells that contain mutant KRAS. Suppression of TBK1 induced apoptosis specifically in human cancer cell lines that depend on oncogenic KRAS expression. In these cells, TBK1 activated NF-kappaB anti-apoptotic signals involving c-Rel and BCL-XL (also known as BCL2L1) that were essential for survival, providing mechanistic insights into this synthetic lethal interaction. These observations indicate that TBK1 and NF-kappaB signalling are essential in KRAS mutant tumours, and establish a general approach for the rational identification of co-dependent pathways in cancer.


Asunto(s)
Genes ras/genética , Proteína Oncogénica p21(ras)/genética , Proteína Oncogénica p21(ras)/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Interferencia de ARN , Alelos , Apoptosis , Línea Celular Tumoral , Supervivencia Celular , Perfilación de la Expresión Génica , Genes Letales , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas c-rel/metabolismo , Transducción de Señal , Proteína bcl-X/metabolismo
17.
Breast Cancer Res ; 16(2): R25, 2014 Mar 14.
Artículo en Inglés | MEDLINE | ID: mdl-24625110

RESUMEN

INTRODUCTION: There is an unmet clinical need for biomarkers to identify breast cancer patients at an increased risk of developing brain metastases. The objective is to identify gene signatures and biological pathways associated with human epidermal growth factor receptor 2-positive (HER2+) brain metastasis. METHODS: We combined laser capture microdissection and gene expression microarrays to analyze malignant epithelium from HER2+ breast cancer brain metastases with that from HER2+ nonmetastatic primary tumors. Differential gene expression was performed including gene set enrichment analysis (GSEA) using publicly available breast cancer gene expression data sets. RESULTS: In a cohort of HER2+ breast cancer brain metastases, we identified a gene expression signature that anti-correlates with overexpression of BRCA1. Sequence analysis of the HER2+ brain metastases revealed no pathogenic mutations of BRCA1, and therefore the aforementioned signature was designated BRCA1 Deficient-Like (BD-L). Evaluation of an independent cohort of breast cancer metastases demonstrated that BD-L values are significantly higher in brain metastases as compared to other metastatic sites. Although the BD-L signature is present in all subtypes of breast cancer, it is significantly higher in BRCA1 mutant primary tumors as compared with sporadic breast tumors. Additionally, BD-L signature values are significantly higher in HER2-/ER- primary tumors as compared with HER2+/ER + and HER2-/ER + tumors. The BD-L signature correlates with breast cancer cell line pharmacologic response to a combination of poly (ADP-ribose) polymerase (PARP) inhibitor and temozolomide, and the signature outperformed four published gene signatures of BRCA1/2 deficiency. CONCLUSIONS: A BD-L signature is enriched in HER2+ breast cancer brain metastases without pathogenic BRCA1 mutations. Unexpectedly, elevated BD-L values are found in a subset of primary tumors across all breast cancer subtypes. Evaluation of pharmacological sensitivity in breast cancer cell lines representing all breast cancer subtypes suggests the BD-L signature may serve as a biomarker to identify sporadic breast cancer patients who might benefit from a therapeutic combination of PARP inhibitor and temozolomide and may be indicative of a dysfunctional BRCA1-associated pathway.


Asunto(s)
Proteína BRCA1/genética , Neoplasias Encefálicas/genética , Neoplasias de la Mama/genética , Regulación Neoplásica de la Expresión Génica , Transcriptoma , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/secundario , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Línea Celular , Línea Celular Tumoral , Daño del ADN , Análisis Mutacional de ADN , Dacarbazina/administración & dosificación , Dacarbazina/análogos & derivados , Inhibidores Enzimáticos/administración & dosificación , Inhibidores Enzimáticos/farmacología , Femenino , Humanos , Inmunohistoquímica , Análisis de Secuencia por Matrices de Oligonucleótidos , Ftalazinas/administración & dosificación , Piperazinas/administración & dosificación , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Poli(ADP-Ribosa) Polimerasas/genética , Poli(ADP-Ribosa) Polimerasas/metabolismo , Pronóstico , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Temozolomida
18.
Proc Natl Acad Sci U S A ; 108(31): 12845-50, 2011 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-21757645

RESUMEN

Human tumors often contain slowly proliferating cancer cells that resist treatment, but we do not know precisely how these cells arise. We show that rapidly proliferating cancer cells can divide asymmetrically to produce slowly proliferating "G0-like" progeny that are enriched following chemotherapy in breast cancer patients. Asymmetric cancer cell division results from asymmetric suppression of AKT/PKB kinase signaling in one daughter cell during telophase of mitosis. Moreover, inhibition of AKT signaling with small-molecule drugs can induce asymmetric cancer cell division and the production of slow proliferators. Cancer cells therefore appear to continuously flux between symmetric and asymmetric division depending on the precise state of their AKT signaling network. This model may have significant implications for understanding how tumors grow, evade treatment, and recur.


Asunto(s)
División Celular , Neoplasias/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Western Blotting , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Células HCT116 , Compuestos Heterocíclicos con 3 Anillos/química , Compuestos Heterocíclicos con 3 Anillos/farmacología , Humanos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Confocal , Modelos Biológicos , Estructura Molecular , Neoplasias/genética , Neoplasias/patología , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/genética , Fase de Descanso del Ciclo Celular , Transducción de Señal/efectos de los fármacos , Factores de Tiempo
19.
bioRxiv ; 2024 Mar 14.
Artículo en Inglés | MEDLINE | ID: mdl-38559183

RESUMEN

Circulating Tumor Cells (CTCs), interrogated by sampling blood from patients with cancer, contain multiple analytes, including intact RNA, high molecular weight DNA, proteins, and metabolic markers. However, the clinical utility of tumor cell-based liquid biopsy has been limited since CTCs are very rare, and current technologies cannot process the blood volumes required to isolate a sufficient number of tumor cells for in-depth assays. We previously described a high-throughput microfluidic prototype utilizing high-flow channels and amplification of cell sorting forces through magnetic lenses. Here, we apply this technology to analyze patient-derived leukapheresis products, interrogating a mean blood volume of 5.83 liters from patients with metastatic cancer, with a median of 2,799 CTCs purified per patient. Isolation of many CTCs from individual patients enables characterization of their morphological and molecular heterogeneity, including cell and nuclear size and RNA expression. It also allows robust detection of gene copy number variation, a definitive cancer marker with potential diagnostic applications. High-volume microfluidic enrichment of CTCs constitutes a new dimension in liquid biopsies.

20.
Nat Methods ; 7(8): 619-21, 2010 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-20639869

RESUMEN

Generating reliable expression profiles from minute cell quantities is critical for scientific discovery and potential clinical applications. Here we present low-quantity digital gene expression (LQ-DGE), an amplification-free approach involving capture of poly(A)(+) RNAs from cellular lysates onto poly(dT)-coated sequencing surfaces, followed by on-surface reverse transcription and sequencing. We applied LQ-DGE to profile malignant and nonmalignant mouse and human cells, demonstrating its quantitative power and potential applicability to archival specimens.


Asunto(s)
Perfilación de la Expresión Génica/métodos , Animales , Línea Celular , Humanos , Ratones , Análisis por Micromatrices/métodos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Poli T/metabolismo , ARN Mensajero/metabolismo , Transcripción Reversa , Análisis de Secuencia de ADN
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