title: miR-34a predicts survival of Ewing's sarcoma patients and directly influences cell chemo-sensitivity and malignancy.
abstract: Identification of factors to detect chemotherapy-resistant tumours at diagnosis is a first priority for risk-adapted therapy in the oncology of children and young adults, where more individualized, effective, and less toxic treatments are highly desirable. In this study, we analysed the miRNAs discriminating Ewing's sarcoma (EWS) patients with different clinical outcomes in order to identify new indicators of prognosis. miRNA expression was investigated in 49 primary EWSs by using the Agilent human miRNA microarray v.2 and/or qRT-PCR. Statistical power of the samples studied for miRNA expression was verified, indicating adequate sample size. Microarray analysis defined a signature of five miRNAs (miR-34a, miR-23a, miR-92a, miR-490-3p, and miR-130b) as an independent predictor of risk for disease progression and survival. Validation analysis in the extended sample set indicated that both miR-34a and miR-490-3p achieved sufficient statistical power to predict prognosis. Results were particularly robust for miR-34a, which appeared associated with either event-free or overall survival and emerged as a significant predictor also after multivariate analysis. Patients with the highest expression of miR-34a did not experience adverse events in 5 years; in contrast, patients with the lowest expression recurred within 2 years. High expression of miR34a can be detected also in paraffin-embedded tissues by in situ hybridization, thus contributing to an easy routine evaluation of this miRNA. Functional analysis of miR-34a in EWS cell lines indicated that when miR-34a expression was enforced, cells were less proliferative, less malignant, and sensitized to doxorubicin and vincristine. Expression of miR-34a could be increased in p53wt cells by treatment with nutlin-3a. Accordingly, nutlin-3a synergizes with doxorubicin. Overall, our data indicate that miR-34a expression is a strong predictor of outcome in EWS. Restoration of miR-34a activity may be useful to decrease malignancy and increase tumour sensitivity to current drugs, so sparing excessive long-term toxicity to EWS patients.
authors.affiliation: Laboratory of Experimental Oncology, Rizzoli Institute, Bologna, Italy.
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Nakatani
authors.forename: Manuela
authors.initials: M
authors.lastname: Ferracin
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Manara
authors.forename: Selena
authors.initials: S
authors.lastname: Ventura
authors.forename: Valentina
authors.initials: V
authors.lastname: Del Monaco
authors.forename: Stefano
authors.initials: S
authors.lastname: Ferrari
authors.forename: Marco
authors.initials: M
authors.lastname: Alberghini
authors.forename: Andrea
authors.initials: A
authors.lastname: Grilli
authors.forename: Sakari
authors.initials: S
authors.lastname: Knuutila
authors.forename: Karl-Ludwig
authors.initials: KL
authors.lastname: Schaefer
authors.forename: Gianfranco
authors.initials: G
authors.lastname: Mattia
authors.forename: Massimo
authors.initials: M
authors.lastname: Negrini
authors.forename: Piero
authors.initials: P
authors.lastname: Picci
authors.forename: Massimo
authors.initials: M
authors.lastname: Serra
authors.forename: Katia
authors.initials: K
authors.lastname: Scotlandi
mesh_terms: D000971:Antineoplastic Combined Chemotherapy Protocols / Q000627:therapeutic use*; D014408:Biomarkers, Tumor / Q000378:metabolism*; D001859:Bone Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism* / Q000401:mortality / Q000473:pathology; D045744:Cell Line, Tumor; D018572:Disease-Free Survival; D004305:Dose-Response Relationship, Drug; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D004357:Drug Synergism; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D017403:In Situ Hybridization; D020128:Inhibitory Concentration 50; D053208:Kaplan-Meier Estimate; D035683:MicroRNAs / Q000378:metabolism*; D015999:Multivariate Analysis; D020411:Oligonucleotide Array Sequence Analysis; D016612:Paraffin Embedding; D010879:Piperazines / Q000494:pharmacology; D016016:Proportional Hazards Models; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D012189:Retrospective Studies; D018570:Risk Assessment; D012307:Risk Factors; D012512:Sarcoma, Ewing / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism* / Q000401:mortality / Q000473:pathology; D013997:Time Factors; D014162:Transfection; D016896:Treatment Outcome; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D014750:Vincristine / Q000494:pharmacology
chemical_list: D014408:Biomarkers, Tumor; D007093:Imidazoles; C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D010879:Piperazines; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C482205:nutlin 3; D014750:Vincristine; D004317:Doxorubicin