title: Molecular profiling of circulating tumor cells links plasticity to the metastatic process in endometrial cancer.
abstract: BACKGROUND
  About 20% of patients diagnosed with endometrial cancer (EC) are considered high-risk with unfavorable prognosis. In the framework of the European Network for Individualized Treatment in EC (ENITEC), we investigated the presence and phenotypic features of Circulating Tumor Cells (CTC) in high-risk EC patients.
  
  
  METHODS
  CTC isolation was carried out in peripheral blood samples from 34 patients, ranging from Grade 3 Stage IB to Stage IV carcinomas and recurrences, and 27 healthy controls using two methodologies. Samples were subjected to EpCAM-based immunoisolation using the CELLection™ Epithelial Enrich kit (Invitrogen, Dynal) followed by RTqPCR analysis. The phenotypic determinants of endometrial CTC in terms of pathogenesis, hormone receptor pathways, stem cell markers and epithelial to mesenchymal transition (EMT) drivers were asked. Kruskal-Wallis analysis followed by Dunn's post-test was used for comparisons between groups. Statistical significance was set at p < 0.05.
  
  
  RESULTS
  EpCAM-based immunoisolation positively detected CTC in high-risk endometrial cancer patients. CTC characterization indicated a remarkable plasticity phenotype defined by the expression of the EMT markers ETV5, NOTCH1, SNAI1, TGFB1, ZEB1 and ZEB2. In addition, the expression of ALDH and CD44 pointed to an association with stemness, while the expression of CTNNB1, STS, GDF15, RELA, RUNX1, BRAF and PIK3CA suggested potential therapeutic targets. We further recapitulated the EMT phenotype found in endometrial CTC through the up-regulation of ETV5 in an EC cell line, and validated in an animal model of systemic dissemination the propensity of these CTC in the accomplishment of metastasis.
  
  
  CONCLUSIONS
  Our results associate the presence of CTC with high-risk EC. Gene-expression profiling characterized a CTC-plasticity phenotype with stemness and EMT features. We finally recapitulated this CTC-phenotype by over-expressing ETV5 in the EC cell line Hec1A and demonstrated an advantage in the promotion of metastasis in an in vivo mouse model of CTC dissemination and homing.
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authors.lastname: Alonso-Alconada
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authors.forename: Amparo
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authors.lastname: Cano
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authors.lastname: Gil-Moreno
authors.forename: Luis
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authors.lastname: Chiva
authors.forename: Juan
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authors.lastname: Cueva
authors.forename: Maria
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authors.lastname: Vieito
authors.forename: Eugenia
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authors.lastname: Ortega
authors.forename: Javier
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authors.lastname: Mariscal
authors.forename: Eva
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authors.lastname: Colas
authors.forename: Josep
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authors.lastname: Castellvi
authors.forename: Maite
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authors.lastname: Cusido
authors.forename: Xavier
authors.initials: X
authors.lastname: Dolcet
authors.forename: Hans W
authors.initials: HW
authors.lastname: Nijman
authors.forename: Tjalling
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authors.lastname: Bosse
authors.forename: John A
authors.initials: JA
authors.lastname: Green
authors.forename: Andrea
authors.initials: A
authors.lastname: Romano
authors.forename: Jaume
authors.initials: J
authors.lastname: Reventos
authors.forename: Rafael
authors.initials: R
authors.lastname: Lopez-Lopez
authors.forename: Helga B
authors.initials: HB
authors.lastname: Salvesen
authors.forename: Frederic
authors.initials: F
authors.lastname: Amant
authors.forename: Xavier
authors.initials: X
authors.lastname: Matias-Guiu
authors.forename: Gema
authors.initials: G
authors.lastname: Moreno-Bueno
authors.affiliation: Translational Medical Oncology; Health Research Institute of Santiago (IDIS), SERGAS, Trav, Choupana s/n 15706, Santiago de Compostela, Spain. miguel.abal.posada@sergas.es.
authors.forename: Miguel
authors.initials: M
authors.lastname: Abal
mesh_terms: D000368:Aged; D000818:Animals; D002469:Cell Separation; D004268:DNA-Binding Proteins / Q000378:metabolism; D004195:Disease Models, Animal; D016889:Endometrial Neoplasms / Q000235:genetics* / Q000473:pathology*; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008819:Mice, Nude; D009362:Neoplasm Metastasis; D009360:Neoplastic Cells, Circulating / Q000378:metabolism* / Q000473:pathology*; D010641:Phenotype; D012307:Risk Factors; D014157:Transcription Factors / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; C086680:ETV5 protein, human; D014157:Transcription Factors