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1.
Future Oncol ; 16(28): 2177-2189, 2020 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-32716216

RESUMEN

Aim: To determine the concordance between plasma and tissue RAS mutation status in metastatic colorectal cancer patients to gauge whether blood-based testing is a viable alternative. We also evaluated the change in mutation status on progression. Materials/methods: RAS testing was performed on plasma from patients commencing first-line therapy (OncoBEAM™ RAS CEIVD kit). Results were then compared with formalin-fixed paraffin embedded tumor samples. Results: The overall percentage agreement (concordance) was 86.0% (86/100), which demonstrates that blood-based testing is an alternative to tissue-based testing. Reproducibility was 100% between three laboratories and 20% showed changes in their RAS mutational status on progression. Conclusion: These results show good concordance between tissue and plasma samples and suggest the need for longitudinal plasma testing during treatment to guide management decisions.


Asunto(s)
Biomarcadores de Tumor , Genes ras , Mutación , Neoplasias/diagnóstico , Neoplasias/genética , Adulto , Anciano , Anciano de 80 o más Años , Alelos , ADN Tumoral Circulante , Análisis Mutacional de ADN/métodos , Análisis Mutacional de ADN/normas , Progresión de la Enfermedad , Femenino , Humanos , Biopsia Líquida/métodos , Biopsia Líquida/normas , Masculino , Persona de Mediana Edad , Metástasis de la Neoplasia , Estadificación de Neoplasias , Neoplasias/sangre , Neoplasias/terapia , Tiempo de Tratamiento
2.
Oncotarget ; 7(39): 63730-63746, 2016 Sep 27.
Artículo en Inglés | MEDLINE | ID: mdl-27572314

RESUMEN

We analyzed the molecular basis for carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1)-controlled inhibition of epithelial-mesenchymal transition (EMT) in a mouse model for mammary adenocarcinoma (WAP-T mice). We demonstrate that silencing of CEACAM1 in WAP-T tumor-derived G-2 cells induces epithelial-mesenchymal plasticity (EMP), as evidenced by typical changes of gene expression, morphology and increased invasion. In contrast, reintroduction of CEACAM1 into G-2 cells reversed up-regulation of genes imposing mesenchymal transition, as well as cellular invasion. We identified the Wnt-pathway as target for CEACAM1-mediated repression of EMT. Importantly, ß-catenin phosphorylation status and transcriptional activity strongly depend on CEACAM1 expression: CEACAM1high G-2 cells displayed enhanced phosphorylation of ß-catenin at S33/S37/T41 and decreased phosphorylation at Y86, thereby inhibiting canonical Wnt/ß-catenin signaling. We identified Src-homology 2 domain-containing phosphatase 2 (SHP-2) as a critical binding partner of CEACAM1 that could modulate ß-catenin Y86 phosphorylation. Hence, CEACAM1 serves as a scaffold that controls membrane proximal ß-catenin signaling. In vivo, mammary tumors of WAP-T/CEACAM1null mice displayed increased nuclear translocation of ß-catenin and a dramatically enhanced metastasis rate compared to WAP-T mice. Hence, CEACAM1 controls EMT in vitro and in vivo by site-specific regulation of ß-catenin phosphorylation. Survival analyses of human mammary carcinoma patients corroborated these data, indicating that CEACAM1 is a prognostic marker for breast cancer survival.


Asunto(s)
Neoplasias de la Mama/metabolismo , Antígeno Carcinoembrionario/metabolismo , Carcinoma/metabolismo , Transición Epitelial-Mesenquimal , Regulación Neoplásica de la Expresión Génica , Neoplasias Mamarias Experimentales/metabolismo , Animales , Antígenos CD/metabolismo , Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/patología , Moléculas de Adhesión Celular/metabolismo , Línea Celular Tumoral , Femenino , Humanos , Técnicas In Vitro , Neoplasias Mamarias Experimentales/patología , Ratones , Invasividad Neoplásica , Metástasis de la Neoplasia , Fenotipo , Fosforilación , Proteína Tirosina Fosfatasa no Receptora Tipo 11/metabolismo , Transducción de Señal , Regulación hacia Arriba , beta Catenina/metabolismo
3.
BMC Cancer ; 15: 178, 2015 Mar 26.
Artículo en Inglés | MEDLINE | ID: mdl-25886487

RESUMEN

BACKGROUND: Experimental analysis of the metastatic cascade requires suitable model systems which allow tracing of disseminated tumor cells and the identification of factors leading to metastatic outgrowth in distant organs. Such models, especially models using immune-competent mice, are rather scarce. We here analyze tumor cell dissemination and metastasis in an immune-competent transplantable mouse mammary tumor model, based on the SV40 transgenic WAP-T mouse mammary carcinoma model. METHODS: We orthotopically transplanted into immune-competent WAP-T mice two tumor cell lines (H8N8, moderately metastatic, and G-2, non-metastatic), developed from primary WAP-T tumors. G-2 and H8N8 cells exhibit stem cell characteristics, form homeostatic, heterotypic tumor cell systems in vitro, and closely mimic endogenous primary tumors after orthotopic transplantation into syngeneic, immune-competent WAP-T mice. Tumor cell transgene-specific PCR allows monitoring of tumor cell dissemination into distinct organs, and immunohistochemistry for SV40 T-antigen tracing of single disseminated tumor cells (DTC). RESULTS: While only H8N8 cell-derived tumors developed metastases, tumors induced with both cell lines disseminated into a variety of organs with similar efficiency and similar organ distribution. H8N8 metastases arose only in lungs, indicating that organ-specific metastatic outgrowth depends on the ability of DTC to re-establish a tumor cell system rather than on invasion per se. Resection of small tumors (0.5 cm(3)) prevented metastasis of H8N8-derived tumors, most likely due to the rather short half-life of DTC, and thus to shorter exposure of the mice to DTC. In experimental metastasis by tail vein injection, G-2 and H8N8 cells both were able to form lung metastases with similar efficiency. However, after injection of sorted "mesenchymal" and "epithelial" G-2 cell subpopulations, only the "epithelial" subpopulation formed lung metastases. CONCLUSIONS: We demonstrate the utility of our mouse model to analyze factors influencing tumor cell dissemination and metastasis. We suggest that the different metastatic capacity of G-2 and H8N8 cells is due to their different degrees of epithelial-mesenchymal plasticity (EMP), and thus the ability of the respective disseminated cells to revert from a "mesenchymal" to an "epithelial" differentiation state.


Asunto(s)
Neoplasias de la Mama/patología , Carcinogénesis , Neoplasias Mamarias Animales/patología , Metástasis de la Neoplasia/patología , Animales , Antígenos Transformadores de Poliomavirus/genética , Biomarcadores de Tumor/genética , Neoplasias de la Mama/genética , Línea Celular Tumoral , Proliferación Celular/genética , Transición Epitelial-Mesenquimal/genética , Femenino , Humanos , Neoplasias Mamarias Animales/genética , Ratones , Ratones Transgénicos , Metástasis de la Neoplasia/genética
4.
Int J Cancer ; 137(1): 25-36, 2015 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-25449528

RESUMEN

In this study, the effects of the standard chemotherapy, cyclophosphamide/adriamycin/5-fluorouracil (CAF) on tumor growth, dissemination and recurrence after orthotopic implantation of murine G-2 cells were analyzed in the syngeneic immunocompetent whey acidic protein-T mouse model (Wegwitz et al., PLoS One 2010; 5:e12103; Schulze-Garg et al., Oncogene 2000; 19:1028-37). Single-dose CAF treatment reduced tumor size significantly, but was not able to eradicate all tumor cells, as recurrent tumor growth was observed 4 weeks after CAF treatment. Nine days after CAF treatment, residual tumors showed features of regressive alterations and were composed of mesenchymal-like tumor cells, infiltrating immune cells and some tumor-associated fibroblasts with an intense deposition of collagen. Recurrent tumors were characterized by coagulative necrosis and less tumor cell differentiation compared with untreated tumors, suggesting a more aggressive tumor phenotype. In support, tumor cell dissemination was strongly enhanced in mice that had developed recurrent tumors in comparison with untreated controls, although only few disseminated tumor cells could be detected in various organs 9 days after CAF application. In vitro experiments revealed that CAF treatment of G-2 cells eliminates the vast majority of epithelial tumor cells, whereas tumor cells with a mesenchymal phenotype survive. These results together with the in vivo findings suggest that tumor cells that underwent epithelial-mesenchymal transition and/or exhibit stem-cell-like properties are difficult to eliminate using one round of CAF chemotherapy. The model system described here provides a valuable tool for the characterization of the effects of chemotherapeutic regimens on recurrent tumor growth and on tumor cell dissemination, thereby enabling the development and preclinical evaluation of novel therapeutic strategies to target mammary carcinomas.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Neoplasias Mamarias Experimentales/tratamiento farmacológico , Neoplasias Mamarias Experimentales/patología , Recurrencia Local de Neoplasia/patología , Neoplasia Residual/patología , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ciclofosfamida/administración & dosificación , Ciclofosfamida/farmacología , Doxorrubicina/administración & dosificación , Doxorrubicina/farmacología , Transición Epitelial-Mesenquimal/efectos de los fármacos , Femenino , Fluorouracilo/administración & dosificación , Fluorouracilo/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Ratones , Trasplante de Neoplasias
5.
Int J Cancer ; 136(6): E521-33, 2015 Mar 15.
Artículo en Inglés | MEDLINE | ID: mdl-25195563

RESUMEN

To study the postulated mutant p53 (mutp53) "gain of function" effects in mammary tumor development, progression and metastasis, we crossed SV40 transgenic WAP-T mice with mutant p53 transgenic WAP-mutp53 mice. Compared to tumors in monotransgenic WAP-T mice, tumors in bitransgenic WAP-T x WAP-mutp53 mice showed higher tumor grading, enhanced vascularization, and significantly increased metastasis. Bitransgenic tumors revealed a gene signature associated with the oncogenic epithelial-mesenchymal transition pathway (EMT gene signature). In cultures of WAP-T tumor-derived G-2 cancer cells, which are comprised of subpopulations displaying "mesenchymal" and "epithelial" phenotypes, this EMT gene signature was associated with the "mesenchymal" compartment. Furthermore, ectopic expression of mutp53 in G-2 cells sufficed to induce a strong EMT phenotype. In contrast to these in vitro effects, monotransgenic and bitransgenic tumors were phenotypically similar suggesting that in vivo the tumor cell phenotype might be under control of the tumor microenvironment. In support, orthotopic transplantation of G-2 cells as well as of G-2 cells expressing ectopic mutp53 into syngeneic mice resulted in tumors with a predominantly epithelial phenotype, closely similar to that of endogenous primary tumors. We conclude that induction of an EMT gene signature by mutp53 in bitransgenic tumors primarily promotes tumor cell plasticity, that is, the probability of tumor cells to undergo EMT processes under appropriate stimuli, thereby possibly increasing their potential to disseminate and metastasize.


Asunto(s)
Transición Epitelial-Mesenquimal , Neoplasias Mamarias Experimentales/patología , Proteína p53 Supresora de Tumor/fisiología , Animales , Línea Celular Tumoral , Femenino , Humanos , Ratones , Mutación , Metástasis de la Neoplasia , Microambiente Tumoral
6.
Mol Cell Proteomics ; 10(10): M110.006239, 2011 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-21725061

RESUMEN

The Cbl-interacting 85-kDa protein (CIN85) plays an important role as a negative regulator of signaling pathways induced by receptor tyrosine kinases. By assembling multiprotein complexes this versatile adaptor enhances receptor tyrosine kinase-activated clathrin-mediated endocytosis and reduces phosphatidylinositol-3-kinase-induced phosphatidylinositol-3,4,5-trisphosphate production. Here we report the expression of CIN85 in primary splenic B lymphocytes and the B-lymphoma cell lines WEHI 231 and Ba/F3. Cross-linking of the B cell antigen receptor resulted in an increased association of CIN85 with the ubiquitin ligase Cbl. Through a systematic pull-down proteomics approach we identified 51 proteins that interact with CIN85 in B cells, including proteins not shown previously to be CIN85-associated. Among these proteins, the SH2-containing inositol phosphatase 1 (SHIP-1) co-precipitated with both the full-length CIN85 and each of its three SH3 domains. We also showed that this association is constitutive and depends on a region of 79 amino acids near the carboxyl terminus of SHIP-1, a region rich in potential SH3 domain binding sites. Because SHIP-1 is a major negative regulator of the phosphatidylinositol-3-kinase pathway in lymphocytes, we hypothesize that the interaction between SHIP-1 and CIN85 might synergistically facilitate the down-regulation of phosphatidylinositol-3,4,5-trisphosphate levels.


Asunto(s)
Proteínas de Neoplasias/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Fosfatidilinositol 3-Quinasa/metabolismo , Fosfatos de Fosfatidilinositol/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Proteínas Proto-Oncogénicas c-cbl/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Animales , Linfocitos B/metabolismo , Sitios de Unión , Línea Celular Tumoral , Regulación de la Expresión Génica , Humanos , Inositol Polifosfato 5-Fosfatasas , Células Jurkat , Masculino , Espectrometría de Masas , Ratones , Ratones Endogámicos C57BL , Complejos Multiproteicos/metabolismo , Proteínas de Neoplasias/genética , Proteínas del Tejido Nervioso/genética , Fosfatos de Fosfatidilinositol/química , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatasas , Monoéster Fosfórico Hidrolasas/genética , Unión Proteica , Proteómica , Proteínas Recombinantes/metabolismo , Transducción de Señal , Bazo/citología , Dominios Homologos src
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