title: Diagnostic and prognostic microRNAs in the serum of breast cancer patients measured by droplet digital PCR.
abstract: BACKGROUND
  Breast cancer circulating biomarkers include carcinoembryonic antigen and carbohydrate antigen 15-3, which are used for patient follow-up. Since sensitivity and specificity are low, novel and more useful biomarkers are needed. The presence of stable circulating microRNAs (miRNAs) in serum or plasma suggested a promising role for these tiny RNAs as cancer biomarkers. To acquire an absolute concentration of circulating miRNAs and reduce the impact of preanalytical and analytical variables, we used the droplet digital PCR (ddPCR) technique.
  
  
  RESULTS
  We investigated a panel of five miRNAs in the sera of two independent cohorts of breast cancer patients and disease-free controls. The study showed that miR-148b-3p and miR-652-3p levels were significantly lower in the serum of breast cancer patients than that in controls in both cohorts. For these two miRNAs, the stratification of breast cancer patients versus controls was confirmed by receiver operating characteristic curve analyses. In addition, we showed that higher levels of serum miR-10b-5p were associated with clinicobiological markers of poor prognosis.
  
  
  CONCLUSIONS
  The study revealed the usefulness of the ddPCR approach for the quantification of circulating miRNAs. The use of the ddPCR quantitative approach revealed very good agreement between two independent cohorts in terms of comparable absolute miRNA concentrations and consistent trends of dysregulation in breast cancer patients versus controls. Overall, this study supports the use of the quantitative ddPCR approach for monitoring the absolute levels of diagnostic and prognostic tumor-specific circulating miRNAs.
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Mangolini
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy ; Laboratory for Technologies of Advanced Therapies (LTTA), University of Ferrara, Ferrara, Italy.
authors.forename: Manuela
authors.initials: M
authors.lastname: Ferracin
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Maria Vittoria
authors.initials: MV
authors.lastname: Zanzi
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Saccenti
authors.affiliation: Department of Life Sciences and Biotechnology, University of Ferrara, Ferrara, Italy.
authors.forename: Sayda Omer
authors.initials: SO
authors.lastname: Ebnaof
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Valentina Vultaggio
authors.initials: VV
authors.lastname: Poma
authors.affiliation: Department of Medical Sciences, University of Ferrara, Ferrara, Italy.
authors.forename: Juana M
authors.initials: JM
authors.lastname: Sanz
authors.affiliation: Department of Medical Sciences, University of Ferrara, Ferrara, Italy.
authors.forename: Angela
authors.initials: A
authors.lastname: Passaro
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Massimo
authors.initials: M
authors.lastname: Pedriali
authors.affiliation: Oncology Unit, University Hospital of Ferrara, Ferrara, Italy.
authors.forename: Antonio
authors.initials: A
authors.lastname: Frassoldati
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Patrizia
authors.initials: P
authors.lastname: Querzoli
authors.affiliation: Department of Life Sciences and Biotechnology, University of Ferrara, Ferrara, Italy.
authors.forename: Silvia
authors.initials: S
authors.lastname: Sabbioni
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Carcoforo
authors.affiliation: Mercy Cancer Resource Center/Women's Center, Oklahoma City, Oklahoma USA.
authors.forename: Alan
authors.initials: A
authors.lastname: Hollingsworth
authors.affiliation: Department of Morphology, Surgery and Experimental Medicine, University of Ferrara, Via Luigi Borsari, 46, 44121 Ferrara, Italy ; Laboratory for Technologies of Advanced Therapies (LTTA), University of Ferrara, Ferrara, Italy.
authors.forename: Massimo
authors.initials: M
authors.lastname: Negrini
keywords: Breast cancer; Circulating miRNAs; Diagnostic markers; Droplet digital PCR; Prognostic markers