RESUMEN
Copy number gains in genes coding for Rho activating exchange factors as well as losses affecting genes coding for RhoGAP proteins are common in breast cancer (BC), suggesting that elevated Rho signaling may play an important role. Extra copies and overexpression of RHOC also occur, although a role for RhoC overexpression in driving tumor formation has not been assessed in vivo. To this end, we report on the development of a Rosa26 (R26)-targeted Cre-conditional RhoC overexpression mouse (R26RhoC). This mouse was crossed to two models for ERBB2/NEU+ breast cancer: one based on expression of an oncogenic ErbB2/Neu cDNA downstream of the endogenous ErbB2 promoter (FloxNeoNeuNT), the other, a metastatic model that is based on high-level expression from MMTV regulatory elements (NIC). RhoC overexpression dramatically enhanced mammary tumor formation in FloxNeoNeuNT mice but showed a more subtle effect in the NIC line, which forms multiple mammary tumors after a very short latency. RhoC overexpression also enhanced mammary tumor formation in an activated Pik3ca model for breast cancer (Pik3caH1047R). The transforming effect of RhoC was associated with epithelial/mesenchymal transition (EMT) in ErbB2/NeuNT and Pik3caH1047R systems. Thus, our study reveals the importance of elevated wildtype Rho protein expression as a driver of breast tumor formation and highlights the significance of Copy Number Abberations that affect Rho signalling.
Asunto(s)
Fosfatidilinositol 3-Quinasa Clase I , Regulación Neoplásica de la Expresión Génica , Fosfatidilinositol 3-Quinasas , Receptor ErbB-2 , Proteínas de Unión al GTP rho , Proteína rhoC de Unión a GTP , Animales , Receptor ErbB-2/metabolismo , Receptor ErbB-2/genética , Femenino , Proteína rhoC de Unión a GTP/metabolismo , Proteína rhoC de Unión a GTP/genética , Ratones , Fosfatidilinositol 3-Quinasa Clase I/genética , Fosfatidilinositol 3-Quinasa Clase I/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasas/genética , Proteínas de Unión al GTP rho/metabolismo , Proteínas de Unión al GTP rho/genética , Humanos , Ratones Transgénicos , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/patología , Neoplasias Mamarias Experimentales/metabolismo , Transición Epitelial-Mesenquimal/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Transducción de SeñalRESUMEN
The most common events in breast cancer (BC) involve chromosome arm losses and gains. Here we describe identification of 1089 gene-centric common insertion sites (gCIS) from transposon-based screens in 8 mouse models of BC. Some gCIS are driver-specific, others driver non-specific, and still others associated with tumor histology. Processes affected by driver-specific and histology-specific mutations include well-known cancer pathways. Driver non-specific gCIS target the Mediator complex, Ca++ signaling, Cyclin D turnover, RNA-metabolism among other processes. Most gCIS show single allele disruption and many map to genomic regions showing high-frequency hemizygous loss in human BC. Two gCIS, Nf1 and Trps1, show synthetic haploinsufficient tumor suppressor activity. Many gCIS act on the same pathway responsible for tumor initiation, thereby selecting and sculpting just enough and just right signaling. These data highlight ~1000 genes with predicted conditional haploinsufficient tumor suppressor function and the potential to promote chromosome arm loss in BC.
Asunto(s)
Neoplasias de la Mama/genética , Pérdida de Heterocigocidad/genética , Animales , Neoplasias de la Mama/patología , Transformación Celular Neoplásica , Elementos Transponibles de ADN/genética , Femenino , Genes Supresores de Tumor , Humanos , Ratones , Mutagénesis Insercional , Neoplasias Experimentales , Transducción de SeñalRESUMEN
CDH1 and PIK3CA are the two most frequently mutated genes in invasive lobular carcinoma (ILC) of the breast. Transcription profiling has identified molecular subtypes for ILC, one of which, immune-related (IR), is associated with gene expression linked to lymphocyte and macrophage infiltration. Here, we report that deletion of Cdh1, together with activation of Pik3ca in mammary epithelium of genetically modified mice, leads to formation of IR-ILC-like tumors with immune cell infiltration, as well as gene expression linked to T-regulatory (Treg) cell signaling and activation of targetable immune checkpoint pathways. Interestingly, these tumors show enhanced Rac1- and Yap-dependent transcription and signaling, as well as sensitivity to PI3K, Rac1, and Yap inhibitors in culture. Finally, high-dimensional immunophenotyping in control mouse mammary gland and IR-ILC tumors by mass cytometry shows dramatic alterations in myeloid and lymphoid populations associated with immune suppression and exhaustion, highlighting the potential for therapeutic intervention via immune checkpoint regulators.
Asunto(s)
Cadherinas/fisiología , Carcinoma Lobular/patología , Regulación Neoplásica de la Expresión Génica , Neoplasias Mamarias Animales/patología , Mutación , Fosfatidilinositol 3-Quinasas/fisiología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Carcinoma Lobular/inmunología , Carcinoma Lobular/metabolismo , Proteínas de Ciclo Celular/metabolismo , Fosfatidilinositol 3-Quinasa Clase I , Femenino , Neoplasias Mamarias Animales/inmunología , Neoplasias Mamarias Animales/metabolismo , Ratones , Ratones Noqueados , Células Mieloides/inmunología , Células Mieloides/metabolismo , Células Mieloides/patología , Invasividad Neoplásica , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/patología , Transcriptoma , Proteínas Señalizadoras YAP , Proteínas de Unión al GTP rac/metabolismoRESUMEN
Blood vessels are formed through vasculogenesis, followed by remodeling of the endothelial network through angiogenesis. Many events that occur during embryonic vascular development are recapitulated during adult neoangiogenesis, which is critical to tumor growth and metastasis. Current antiangiogenic tumor therapies, based largely on targeting the vascular endothelial growth factor pathway, show limited clinical benefits, thus necessitating the discovery of alternative targets. Here we report the development of a robust embryonic stem cell-based vascular differentiation assay amenable to small-molecule screens to identify novel modulators of angiogenesis. In this context, RSK and TTK were identified as angiogenic modulators. Inhibition of these pathways inhibited angiogenesis in embryoid bodies and human umbilical vein endothelial cells. Furthermore, inhibition of RSK and TTK reduced tumor growth, vascular density, and improved survival in an in vivo Lewis lung carcinoma mouse model. Our study suggests that RSK and TTK are potential targets for antiangiogenic therapy, and provides an assay system for further pathway screens.
Asunto(s)
Vasos Sanguíneos/embriología , Vasos Sanguíneos/metabolismo , Proteínas de Ciclo Celular/metabolismo , Diferenciación Celular , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Proteínas Quinasas S6 Ribosómicas/metabolismo , Inhibidores de la Angiogénesis/farmacología , Animales , Proteínas de Ciclo Celular/antagonistas & inhibidores , Línea Celular , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Descubrimiento de Drogas , Femenino , Humanos , Ratones , Morfogénesis , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Neovascularización Patológica , Neovascularización Fisiológica/efectos de los fármacos , Neovascularización Fisiológica/genética , Organogénesis , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Quinasas S6 Ribosómicas/antagonistas & inhibidoresRESUMEN
Breast cancer is associated with alterations in a number of growth factor and hormone-regulated signaling pathways. Mouse models of metastatic breast cancer typically feature mutated oncoproteins that activate PI3K, Stat3, and Ras signaling, but the individual and combined roles of these pathways in breast cancer progression are poorly understood. In this study, we examined the relationship between oncogenic pathway activation and breast cancer subtype by analyzing mouse mammary tumor formation in which each pathway was activated singly or pairwise. All three oncogenes showed cooperation during primary tumor formation, but efficient dissemination was only dependent on Ras. In addition, transcriptional profiling demonstrated that Ras induced adenocarcinomas with molecular characteristics related to human basal-like and HER2(+) tumors. In contrast, Ras combined with PIK3CA(H1047R), an oncogenic mutant linked to ERα(+)/luminal breast cancer in humans, induced metastatic luminal B-like tumors. Consistent with these data, elevated Ras signaling was associated with basal-like and HER2(+) subtype tumors in humans and showed a statistically significant negative association with estrogen receptor (ER) signaling across all breast cancer. Despite this, there are luminal tumors with elevated Ras signaling. Importantly, when considered as a continuous variable, Ras pathway activation was strongly linked to reduced survival of patients with ERα(+) disease independent of PI3K or Stat3 activation. Therefore, our studies suggest that Ras activation is a key determinant for dissemination and poor prognosis of ERα(+)/luminal breast cancer in humans, and hormone therapy supplemented with Ras-targeting agents may be beneficial for treating this aggressive subtype.
Asunto(s)
Neoplasias de la Mama/patología , Invasividad Neoplásica/patología , Transducción de Señal/fisiología , Proteínas ras/metabolismo , Animales , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/mortalidad , Supervivencia sin Enfermedad , Femenino , Inmunohistoquímica , Estimación de Kaplan-Meier , Ratones , Recurrencia Local de Neoplasia/metabolismo , Recurrencia Local de Neoplasia/patología , Análisis de Secuencia por Matrices de OligonucleótidosRESUMEN
BACKGROUND: The Notch signaling pathway plays complex roles in developing lungs, including regulation of proximodistal fates, airway cell specification and differentiation. However, the specific Notch-mediated signals involved in lung development remain unclear. RESULTS: Here we report that Jagged1 is expressed in a subset of bronchial and bronchiolar epithelial cells, where it controls proximal airway cell fate and differentiation. In agreement with previous studies involving disruption of all Notch signaling, we found that deletion of Jagged1 in airway epithelium increased the number of ciliated cells at the expense of Clara cells, a phenotype associated with downregulation of Hes1. Deletion of Jagged1 also led to an increased number of pulmonary neuroendocrine cells (PNEC), suggesting that Jagged1/Notch signaling inhibits PNEC cell fate. As expected, Jagged1 deletion did not affect alveolar cell differentiation, although alveolar septation was impaired, likely an indirect effect of proximal airway defects. Finally, in the postnatal lung, Jagged1 deletion induced mucous metaplasia, accompanied by downregulation of Hes1 and Hes5. CONCLUSIONS: Our results demonstrate that Jagged1-mediated Notch signaling regulates multiple cell fate decisions as well as differentiation in the respiratory system to coordinate lung development and to maintain a balance of airway cell types in adult life.
Asunto(s)
Proteínas de Unión al Calcio/fisiología , Linaje de la Célula , Regulación del Desarrollo de la Expresión Génica , Péptidos y Proteínas de Señalización Intercelular/fisiología , Pulmón/embriología , Proteínas de la Membrana/fisiología , Receptores Notch/metabolismo , Animales , Proteínas de Unión al Calcio/genética , Diferenciación Celular , Células Epiteliales/metabolismo , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intercelular/genética , Proteína Jagged-1 , Pulmón/metabolismo , Proteínas de la Membrana/genética , Ratones , Ratones Transgénicos , Células Neuroendocrinas/metabolismo , Fenotipo , Proteínas Serrate-Jagged , Transducción de Señal , Factores de TiempoRESUMEN
Basal-like breast cancers (BLBC) express a luminal progenitor gene signature. Notch receptor signaling promotes luminal cell fate specification in the mammary gland, while suppressing stem cell self-renewal. Here we show that deletion of Lfng, a sugar transferase that prevents Notch activation by Jagged ligands, enhances stem/progenitor cell proliferation. Mammary-specific deletion of Lfng induces basal-like and claudin-low tumors with accumulation of Notch intracellular domain fragments, increased expression of proliferation-associated Notch targets, amplification of the Met/Caveolin locus, and elevated Met and Igf-1R signaling. Human BL breast tumors, commonly associated with JAGGED expression, elevated MET signaling, and CAVEOLIN accumulation, express low levels of LFNG. Thus, reduced LFNG expression facilitates JAG/NOTCH luminal progenitor signaling and cooperates with MET/CAVEOLIN basal-type signaling to promote BLBC.
Asunto(s)
Neoplasias de la Mama/enzimología , Caveolinas/metabolismo , Transformación Celular Neoplásica/metabolismo , Glicosiltransferasas/metabolismo , Glándulas Mamarias Animales/enzimología , Neoplasias Mamarias Experimentales/enzimología , Células Madre Neoplásicas/enzimología , Proteínas Proto-Oncogénicas c-met/metabolismo , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Proteínas de Unión al Calcio/metabolismo , Caveolinas/genética , Proliferación Celular , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/patología , Células Cultivadas , Claudinas/metabolismo , Bases de Datos Genéticas , Femenino , Perfilación de la Expresión Génica/métodos , Regulación del Desarrollo de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Glicosiltransferasas/deficiencia , Glicosiltransferasas/genética , Humanos , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Proteína Jagged-1 , Glándulas Mamarias Animales/crecimiento & desarrollo , Glándulas Mamarias Animales/patología , Glándulas Mamarias Animales/trasplante , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/patología , Proteínas de la Membrana/metabolismo , Ratones , Ratones Noqueados , Persona de Mediana Edad , Células Madre Neoplásicas/patología , Células Madre Neoplásicas/trasplante , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas c-met/genética , Receptor IGF Tipo 1/metabolismo , Receptores Notch/metabolismo , Proteínas Serrate-Jagged , Transducción de SeñalRESUMEN
PIK3CA, which codes for the p110α catalytic subunit of phosphatidylinositol 3-kinase, is one of the most frequently mutated genes in human breast cancer. Here, we describe a mouse model for PIK3CA-induced breast cancer by using the ROSA26 (R26) knock-in system, in which targeted Pik3ca alleles can be activated through transgenic expression of Cre recombinase. We mated Pik3ca(H1047R) and Pik3ca(wt) knock-in lines with MMTV-Cre transgenics, which express Cre in mammary epithelium. Starting at approximately 5 months of age, female R26-Pik3ca(H1047R);MMTV-Cre mice, but not control R26-Pik3ca(wt);MMTV-Cre mice, developed mammary tumors, as well as lymphoid and skin malignancies. R26-Pik3ca(H1047R);MMTV-Cre mammary tumors were typically either adenosquamous carcinoma or adenomyoepithelioma. As p53 is the most commonly mutated gene in breast cancer, we tested for genetic interaction between Pik3ca(H1047R) and p53 loss-of-function mutations in R26-Pik3ca(H1047R);p53(loxP/+);MMTV-Cre mice. This led to decreased survival of double-mutant animals, which developed lymphoma and mammary tumors with rapid kinetics. Mammary tumors that formed in p53(loxP/+);MMTV-Cre conditional mutants were either poorly differentiated adenocarcinoma or spindle cell/EMT, whereas R26-Pik3ca(H1047R);p53(loxP/+);MMTV-Cre mammary tumors were mostly adenosquamous carcinoma or spindle cell/EMT indicating that double-mutant mice develop a distinct spectrum of mammary tumors. Thus, an oncogenic variant of PIK3CA implicated in multiple human breast cancer subtypes can induce a very diverse spectrum of mammary tumors in mice. Furthermore, Pik3ca(H1047R) shows cooperation with p53, which altered the specific tumors that formed. Thus, the two most frequently mutated genes in human breast cancer show cooperation in mammary tumor formation.