RESUMEN
Epithelial-to-mesenchymal transition (EMT) contributes significantly to chemotherapy resistance and remains a critical challenge in treating advanced breast cancer. The complexity of EMT, involving redundant pro-EMT signaling pathways and its paradox reversal process, mesenchymal-to-epithelial transition (MET), has hindered the development of effective treatments. In this study, we utilized a Tri-PyMT EMT lineage-tracing model and single-cell RNA sequencing (scRNA-seq) to comprehensively analyze the EMT status of tumor cells. Our findings revealed elevated ribosome biogenesis (RiBi) during the transitioning phases of both EMT and MET processes. RiBi and its subsequent nascent protein synthesis mediated by ERK and mTOR signalings are essential for EMT/MET completion. Importantly, inhibiting excessive RiBi genetically or pharmacologically impaired the EMT/MET capability of tumor cells. Combining RiBi inhibition with chemotherapy drugs synergistically reduced metastatic outgrowth of epithelial and mesenchymal tumor cells under chemotherapies. Our study suggests that targeting the RiBi pathway presents a promising strategy for treating patients with advanced breast cancer. Significance: This study uncovers the crucial involvement of ribosome biogenesis (RiBi) in the regulation of epithelial and mesenchymal state oscillations in breast cancer cells, which plays a major role in the development of chemoresistant metastasis. By proposing a novel therapeutic strategy targeting the RiBi pathway, the study offers significant potential to enhance treatment efficacy and outcomes for patients with advanced breast cancer. This approach could help overcome the limitations of current chemotherapy options and address the complex challenges posed by EMT-mediated chemoresistance.
RESUMEN
Radiation therapy (RT) in combination with immune checkpoint inhibitor (ICI) represents a promising regimen for non-small cell lung cancer (NSCLC), however, the underlying mechanisms are poorly characterized. We identified a specific dose of RT that conferred tumor regression and improved survival in NSCLC models when combined with ICI. The immune-modulating functions of RT was ascribed to activated lung-resident Scgb1a1+ club cells. Importantly, mice with club cell-specific knockout of synaptosome-associated protein 23 failed to benefit from the combination treatment, indicating a pivotal role of club cell secretome. We identified 8 club cells secretory proteins, which inhibited immunosuppressive myeloid cells, reduced pro-tumor inflammation, and enhanced anti-tumor immunity. Notably, CC10, a member of club cell secretome was increased in plasma of NSCLC patients responding to the combination therapy. By revealing an immune-regulatory role of club cells, our studies have the potential to guide future clinical trials of ICI in NSCLC.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Animales , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Neoplasias Pulmonares/tratamiento farmacológico , Ratones , Uteroglobina/uso terapéuticoRESUMEN
Copper serves as a co-factor for a host of metalloenzymes that contribute to malignant progression. The orally bioavailable copper chelating agent tetrathiomolybdate (TM) has been associated with a significant survival benefit in high-risk triple negative breast cancer (TNBC) patients. Despite these promising data, the mechanisms by which copper depletion impacts metastasis are poorly understood and this remains a major barrier to advancing TM to a randomized phase II trial. Here, using two independent TNBC models, we report a discrete subpopulation of highly metastatic SOX2/OCT4+ cells within primary tumors that exhibit elevated intracellular copper levels and a marked sensitivity to TM. Global proteomic and metabolomic profiling identifies TM-mediated inactivation of Complex IV as the primary metabolic defect in the SOX2/OCT4+ cell population. We also identify AMPK/mTORC1 energy sensor as an important downstream pathway and show that AMPK inhibition rescues TM-mediated loss of invasion. Furthermore, loss of the mitochondria-specific copper chaperone, COX17, restricts copper deficiency to mitochondria and phenocopies TM-mediated alterations. These findings identify a copper-metabolism-metastasis axis with potential to enrich patient populations in next-generation therapeutic trials.
Asunto(s)
Cobre/metabolismo , Mitocondrias/metabolismo , Neoplasias de la Mama Triple Negativas/metabolismo , Animales , Línea Celular Tumoral , Proteínas Transportadoras de Cobre/genética , Proteínas Transportadoras de Cobre/metabolismo , Femenino , Humanos , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones , Ratones Endogámicos C57BL , Mitocondrias/genética , Metástasis de la Neoplasia , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Fosforilación Oxidativa , Factores de Transcripción SOXB1/genética , Factores de Transcripción SOXB1/metabolismo , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/patologíaRESUMEN
Metastases are responsible for the majority of breast cancer-associated deaths. The contribution of epithelial-to-mesenchymal transition (EMT) in the establishment of metastases is still controversial. To obtain in vivo evidence of EMT in metastasis, we established an EMT lineage tracing (Tri-PyMT) model, in which tumor cells undergoing EMT would irreversibly switch their fluorescent marker from RFP+ to GFP+ due to mesenchymal-specific Cre expression. Surprisingly, we found that lung metastases were predominantly derived from the epithelial compartment of breast tumors. However, concerns were raised on the fidelity and sensitivity of RFP-to-GFP switch of this model in reporting EMT of metastatic tumor cells. Here, we evaluated Tri-PyMT cells at the single-cell level using single-cell RNA-sequencing and found that the Tri-PyMT cells exhibited a spectrum of EMT phenotypes, with EMT-related genes concomitantly expressed with the activation of GFP. The fluorescent color switch in these cells precisely marked an unequivocal change in EMT status, defining the pre-EMT and post-EMT compartments within the tumor. Consistently, the pre-EMT cells played dominant roles in metastasis, while the post-EMT cells were supportive in promoting tumor invasion and angiogenesis. Importantly, the post-EMT (GFP+) cells in the Tri-PyMT model were not permanently committed to the mesenchymal phenotype; they were still capable of reverting to the epithelial phenotype and giving rise to secondary tumors, suggesting their persistent EMT plasticity. Our study addressed major concerns with the Tri-PyMT EMT lineage tracing model, which provides us with a powerful tool to investigate the dynamic EMT process in tumor biology. SIGNIFICANCE: These findings confirm the fidelity and sensitivity of the EMT lineage tracing (Tri-PyMT) model and highlight the differential contributions of pre- and post-EMT tumor cells in breast cancer metastasis.See related commentary by Bunz, p. 153.
Asunto(s)
Neoplasias de la Mama , Neoplasias Pulmonares , Línea Celular Tumoral , Transición Epitelial-Mesenquimal , Humanos , FenotipoRESUMEN
Epigenetic changes are increasingly being appreciated as key events in breast cancer progression. However, breast cancer subtype-specific epigenetic regulation remains poorly investigated. Here we report that EZH2 is a leading candidate of epigenetic modulators associated with the TNBC subtype and that it predicts poor overall survival in TNBC patients. We demonstrate that specific pharmacological or genetic inhibition of EZH2 catalytic activity impairs distant metastasis. We further define a specific EZH2high population with enhanced invasion, mammosphere formation, and metastatic potential that exhibits marked sensitivity to EZH2 inhibition. Mechanistically, EZH2 inhibition differentiates EZH2high basal cells to a luminal-like phenotype by derepressing GATA3 and renders them sensitive to endocrine therapy. Furthermore, dissection of human TNBC heterogeneity shows that EZH2high basal-like 1 and mesenchymal subtypes have exquisite sensitivity to EZH2 inhibition compared with the EZH2low luminal androgen receptor subtype. These preclinical findings provide a rationale for clinical development of EZH2 as a targeted therapy against TNBC metastasis.
Asunto(s)
Biocatálisis , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Animales , Secuencia de Bases , Compartimento Celular , Diferenciación Celular , Línea Celular Tumoral , Proliferación Celular , Progresión de la Enfermedad , Proteína Potenciadora del Homólogo Zeste 2/antagonistas & inhibidores , Epigénesis Genética , Femenino , Factor de Transcripción GATA3/metabolismo , Humanos , Ratones Endogámicos BALB C , Ratones SCID , Proteínas Mutantes/metabolismo , Invasividad Neoplásica , Metástasis de la Neoplasia , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Fenotipo , Factores de Transcripción SOXB1/metabolismo , Neoplasias de la Mama Triple Negativas/genéticaRESUMEN
Success of immune checkpoint inhibitors in advanced non-small-cell lung cancer (NSCLC) has invigorated their use in the neoadjuvant setting for early-stage disease. However, the cellular and molecular mechanisms of the early immune responses to therapy remain poorly understood. Through an integrated analysis of early-stage NSCLC patients and a Kras mutant mouse model, we show a prevalent programmed cell death 1/programmed cell death 1 ligand 1 (PD-1/PD-L1) axis exemplified by increased intratumoral PD-1+ T cells and PD-L1 expression. Notably, tumor progression was associated with spatiotemporal modulation of the immune microenvironment with dominant immunosuppressive phenotypes at later phases of tumor growth. Importantly, PD-1 inhibition controlled tumor growth, improved overall survival, and reprogrammed tumor-associated lymphoid and myeloid cells. Depletion of T lymphocyte subsets demonstrated synergistic effects of those populations on PD-1 inhibition of tumor growth. Transcriptome analyses revealed T cell subset-specific alterations corresponding to degree of response to the treatment. These results provide insights into temporal evolution of the phenotypic effects of PD-1/PD-L1 activation and inhibition and motivate targeting of this axis early in lung cancer progression.
Asunto(s)
Antígeno B7-H1/inmunología , Carcinoma de Pulmón de Células no Pequeñas/inmunología , Neoplasias Pulmonares/inmunología , Receptor de Muerte Celular Programada 1/efectos de los fármacos , Receptor de Muerte Celular Programada 1/inmunología , Animales , Anticuerpos Monoclonales/uso terapéutico , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Pulmón de Células no Pequeñas/terapia , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Humanos , Inmunoterapia , Pulmón/patología , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/terapia , Ratones , Ratones Endogámicos C57BL , Proteínas Proto-Oncogénicas p21(ras) , Linfocitos T , Microambiente Tumoral/inmunologíaRESUMEN
Molecularly targeted therapies benefit approximately 15-20% of non-small cell lung cancer (NSCLC) patients carrying specific drug-sensitive mutations. Thus, there is a clinically unmet need for the identification of novel targets for drug development. Here, we performed RNA-deep sequencing to identify altered gene expression between malignant and non-malignant lung tissue. Matrix Metalloproteinase 14 (MMP14), a membrane-bound proteinase, was significantly up-regulated in the tumor epithelial cells and intratumoral myeloid compartments in both mouse and human NSCLC. Overexpression of a soluble dominant negative MMP14 (DN-MMP14) or pharmacological inhibition of MMP14 blocked invasion of lung cancer cells through a collagen I matrix in vitro and reduced tumor incidence in an orthotopic K-RasG12D/+p53-/- mouse model of lung cancer. Additionally, MMP14 activity mediated proteolytic processing and activation of Heparin-Binding EGF-like Growth Factor (HB-EGF), stimulating the EGFR signaling pathway to increase proliferation and tumor growth. This study highlights the potential for development of therapeutic strategies that target MMP14 in NSCLC with particular focus on MMP14-HB-EGF axis.
Asunto(s)
Factor de Crecimiento Similar a EGF de Unión a Heparina/metabolismo , Neoplasias Pulmonares/metabolismo , Metaloproteinasa 14 de la Matriz/metabolismo , Animales , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular , Transformación Celular Neoplásica/metabolismo , Colágeno Tipo I/metabolismo , Modelos Animales de Enfermedad , Receptores ErbB/metabolismo , Xenoinjertos , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Metaloproteinasa 14 de la Matriz/genética , Ratones , Estadificación de Neoplasias , Proteolisis , Transducción de Señal , Carga TumoralRESUMEN
PURPOSE: Bone marrow-derived progenitor cells, including VEGFR2+ endothelial progenitor cells (EPCs) and copper-dependent pathways, model the tumor microenvironment. We hypothesized that copper depletion using tetrathiomolybdate would reduce EPCs in high risk for patients with breast cancer who have relapsed. We investigated the effect of tetrathiomolybdate on the tumor microenvironment in preclinical models. EXPERIMENTAL DESIGN: Patients with stage II triple-negative breast cancer (TNBC), stage III and stage IV without any evidence of disease (NED), received oral tetrathiomolybdate to maintain ceruloplasmin (Cp) between 8 and 17 mg/dL for 2 years or until relapse. Endpoints were effect on EPCs and other biomarkers, safety, event-free (EFS), and overall survival (OS). For laboratory studies, MDA-LM2-luciferase cells were implanted into CB17-SCID mice and treated with tetrathiomolybdate or water. Tumor progression was quantified by bioluminescence imaging (BLI), copper depletion status by Cp oxidase levels, lysyl oxidase (LOX) activity by ELISA, and collagen deposition. RESULTS: Seventy-five patients enrolled; 51 patients completed 2 years (1,396 cycles). Most common grade 3/4 toxicity was neutropenia (3.7%). Lower Cp levels correlated with reduced EPCs (P = 0.002) and LOXL-2 (P < 0.001). Two-year EFS for patients with stage II-III and stage IV NED was 91% and 67%, respectively. For patients with TNBC, EFS was 90% (adjuvant patients) and 69% (stage IV NED patients) at a median follow-up of 6.3 years, respectively. In preclinical models, tetrathiomolybdate decreased metastases to lungs (P = 0.04), LOX activity (P = 0.03), and collagen crosslinking (P = 0.012). CONCLUSIONS: Tetrathiomolybdate is safe, well tolerated, and affects copper-dependent components of the tumor microenvironment. Biomarker-driven clinical trials in high risk for patients with recurrent breast cancer are warranted. Clin Cancer Res; 23(3); 666-76. ©2016 AACR.
Asunto(s)
Adenocarcinoma/secundario , Neoplasias de la Mama/tratamiento farmacológico , Quelantes/uso terapéutico , Cobre/metabolismo , Células Progenitoras Endoteliales/efectos de los fármacos , Neoplasias Pulmonares/secundario , Molibdeno/uso terapéutico , Microambiente Tumoral/efectos de los fármacos , Adenocarcinoma/tratamiento farmacológico , Adenocarcinoma/prevención & control , Aminoácido Oxidorreductasas/sangre , Animales , Neoplasias de la Mama/patología , Línea Celular Tumoral , Ceruloplasmina/análisis , Quelantes/farmacología , Progresión de la Enfermedad , Supervivencia sin Enfermedad , Células Progenitoras Endoteliales/fisiología , Femenino , Estudios de Seguimiento , Humanos , Neoplasias Pulmonares/prevención & control , Ratones SCID , Molibdeno/farmacología , Proteínas de Neoplasias/sangre , Neovascularización Patológica/fisiopatología , Neovascularización Patológica/prevención & control , Neutropenia/inducido químicamente , Riesgo , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Neoplasias de la Mama Triple Negativas/patología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Emerging studies have begun to demonstrate that reprogrammed stromal cells play pivotal roles in tumor growth, metastasis, and resistance to therapy. However, the contribution of stromal cells to non-small-cell lung cancer (NSCLC) has remained underexplored. We used an orthotopic model of Kras-driven NSCLC to systematically dissect the contribution of specific hematopoietic stromal cells in lung cancer. RNA deep-sequencing analysis of individually sorted myeloid lineage and tumor epithelial cells revealed cell-type-specific differentially regulated genes, indicative of activated stroma. We developed a computational model for crosstalk signaling discovery based on ligand-receptor interactions and downstream signaling networks and identified known and novel tumor-stroma paracrine and tumor autocrine crosstalk-signaling pathways in NSCLC. We provide cellular and molecular insights into components of the lung cancer microenvironment that contribute to carcinogenesis. This study has the potential for development of therapeutic strategies that target tumor-stroma interactions and may complement conventional anti-cancer treatments.
Asunto(s)
Perfilación de la Expresión Génica , Células del Estroma/metabolismo , Algoritmos , Animales , Comunicación Autocrina , Células de la Médula Ósea/citología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Células Cultivadas , Modelos Animales de Enfermedad , Células Epiteliales/citología , Células Epiteliales/metabolismo , Interleucina-6/metabolismo , Pulmón/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones , Comunicación Paracrina , Receptores de Interleucina-6/metabolismo , Análisis de Secuencia de ARN , Células del Estroma/citología , Transcriptoma , Microambiente Tumoral , Proteínas ras/genética , Proteínas ras/metabolismoRESUMEN
Tumor endothelial cells (ECs) promote cancer progression in ways beyond their role as conduits supporting metabolism. However, it is unknown how vascular niche-derived paracrine factors, defined as angiocrine factors, provoke tumor aggressiveness. Here, we show that FGF4 produced by B cell lymphoma cells (LCs) through activating FGFR1 upregulates the Notch ligand Jagged1 (Jag1) on neighboring ECs. In turn, upregulation of Jag1 on ECs reciprocally induces Notch2-Hey1 in LCs. This crosstalk enforces aggressive CD44(+)IGF1R(+)CSF1R(+) LC phenotypes, including extranodal invasion and chemoresistance. Inducible EC-selective deletion of Fgfr1 or Jag1 in the Eµ-Myc lymphoma model or impairing Notch2 signaling in mouse and human LCs diminished lymphoma aggressiveness and prolonged mouse survival. Thus, targeting the angiocrine FGF4-FGFR1/Jag1-Notch2 loop inhibits LC aggressiveness and enhances chemosensitivity.
Asunto(s)
Linfoma de Burkitt/metabolismo , Linfoma de Burkitt/patología , Proteínas de Unión al Calcio/metabolismo , Resistencia a Antineoplásicos , Factor 4 de Crecimiento de Fibroblastos/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Proteínas de la Membrana/metabolismo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor Notch2/metabolismo , Animales , Linfoma de Burkitt/genética , Proteínas de Unión al Calcio/genética , Proteínas de Ciclo Celular/metabolismo , Proliferación Celular , Células Endoteliales/metabolismo , Activación Enzimática , Genes myc , Humanos , Receptores de Hialuranos/metabolismo , Péptidos y Proteínas de Señalización Intercelular/genética , Proteína Jagged-1 , Proteínas de la Membrana/genética , Ratones , Ratones Transgénicos , Invasividad Neoplásica , Interferencia de ARN , ARN Interferente Pequeño , Receptor IGF Tipo 1/metabolismo , Receptor de Factor Estimulante de Colonias de Macrófagos/metabolismo , Proteínas Serrate-Jagged , Transducción de Señal/genética , Células Tumorales Cultivadas , Regulación hacia ArribaRESUMEN
UNLABELLED: Metastatic tumors have been shown to establish permissive microenvironments for metastases via recruitment of bone marrow-derived cells. Here, we show that metastasis-incompetent tumors are also capable of generating such microenvironments. However, in these situations, the otherwise prometastatic Gr1(+) myeloid cells create a metastasis-refractory microenvironment via the induction of thrombospondin-1 (Tsp-1) by tumor-secreted prosaposin. Bone marrow-specific genetic deletion of Tsp-1 abolished the inhibition of metastasis, which was restored by bone marrow transplant from Tsp-1(+) donors. We also developed a 5-amino acid peptide from prosaposin as a pharmacologic inducer of Tsp-1 in Gr1(+) bone marrow cells, which dramatically suppressed metastasis. These results provide mechanistic insights into why certain tumors are deficient in metastatic potential and implicate recruited Gr1(+) myeloid cells as the main source of Tsp-1. The results underscore the plasticity of Gr1(+) cells, which, depending on the context, promote or inhibit metastasis, and suggest that the peptide could be a potential therapeutic agent against metastatic cancer. SIGNIFICANCE: The mechanisms of metastasis suppression are poorly understood. Here, we have identified a novel mechanism whereby metastasis-incompetent tumors generate metastasis-suppressive microenvironments in distant organs by inducing Tsp-1 expression in the bone marrowderived Gr1+myeloid cells. A 5-amino acid peptide with Tsp-1inducing activity was identified as a therapeutic agent against metastatic cancer.