title: miR-16-5p Is a Stably-Expressed Housekeeping MicroRNA in Breast Cancer Tissues from Primary Tumors and from Metastatic Sites.
abstract: For quantitative microRNA analyses in formalin-fixed paraffin-embedded (FFPE) tissue, expression levels have to be normalized to endogenous controls. To investigate the most stably-expressed microRNAs in breast cancer and its surrounding tissue, we used tumor samples from primary tumors and from metastatic sites. MiRNA profiling using TaqMan(®) Array Human MicroRNA Cards, enabling quantification of 754 unique human miRNAs, was performed in FFPE specimens from 58 patients with metastatic breast cancer. Forty-two (72%) samples were collected from primary tumors and 16 (28%) from metastases. In a cross-platform analysis of a validation cohort of 32 FFPE samples from patients with early breast cancer genome-wide microRNA expression analysis using SurePrintG3 miRNA (8 × 60 K)(®) microarrays from Agilent(®) was performed. Eleven microRNAs could be detected in all samples analyzed. Based on NormFinder and geNorm stability values and the high correlation (rho ≥ 0.8) with the median of all measured microRNAs, miR-16-5p, miR-29a-3p, miR-126-3p, and miR-222-3p are suitable single gene housekeeper candidates. In the cross-platform validation, 29 human microRNAs were strongly expressed (mean log2-intensity > 10) and 21 of these microRNAs including miR-16-5p and miR-29a-3p were also stably expressed (CV < 5%). Thus, miR-16-5p and miR-29a-3p are both strong housekeeper candidates. Their Normfinder stability values calculated across the primary tumor and metastases subgroup indicate that miR-29a-3p can be considered as the strongest housekeeper in a cohort with mainly samples from primary tumors, whereas miR-16-5p might perform better in a metastatic sample enriched cohort.
authors.affiliation: IIIrd Medical Department with Hematology and Medical Oncology, Oncologic Center, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. g.rinnerthaler@salk.at.|Salzburg Cancer Research Institute with Laboratory of Immunological and Molecular Cancer Research and Center for Clinical Cancer and Immunology Trials, Salzburg 5020, Austria. g.rinnerthaler@salk.at.|Cancer Cluster Salzburg, Salzburg 5020, Austria. g.rinnerthaler@salk.at.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Rinnerthaler
authors.affiliation: Division of Bioinformatics, Biocenter, Medical University of Innsbruck, Innsbruck 6020, Austria. hubert.hackl@i-med.ac.at.
authors.forename: Hubert
authors.initials: H
authors.lastname: Hackl
authors.affiliation: IIIrd Medical Department with Hematology and Medical Oncology, Oncologic Center, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. s.gampenrieder@salk.at.
authors.forename: Simon Peter
authors.initials: SP
authors.lastname: Gampenrieder
authors.affiliation: IIIrd Medical Department with Hematology and Medical Oncology, Oncologic Center, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. Frank.Hamacher71@gmx.de.
authors.forename: Frank
authors.initials: F
authors.lastname: Hamacher
authors.affiliation: IIIrd Medical Department with Hematology and Medical Oncology, Oncologic Center, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. cl.hufnagl@salk.at.
authors.forename: Clemens
authors.initials: C
authors.lastname: Hufnagl
authors.affiliation: Department of Pathology, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. c.kronberger@salk.at.
authors.forename: Cornelia
authors.initials: C
authors.lastname: Hauser-Kronberger
authors.affiliation: Department of Radiotherapy, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. f.zehentmayr@salk.at.
authors.forename: Franz
authors.initials: F
authors.lastname: Zehentmayr
authors.affiliation: Department of Radiotherapy, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. g.fastner@salk.at.
authors.forename: Gerd
authors.initials: G
authors.lastname: Fastner
authors.affiliation: Department of Radiotherapy, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. f.sedlmayer@salk.at.
authors.forename: Felix
authors.initials: F
authors.lastname: Sedlmayer
authors.affiliation: IIIrd Medical Department with Hematology and Medical Oncology, Oncologic Center, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. b.mlineritsch@salk.at.|Salzburg Cancer Research Institute with Laboratory of Immunological and Molecular Cancer Research and Center for Clinical Cancer and Immunology Trials, Salzburg 5020, Austria. b.mlineritsch@salk.at.|Cancer Cluster Salzburg, Salzburg 5020, Austria. b.mlineritsch@salk.at.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Mlineritsch
authors.affiliation: IIIrd Medical Department with Hematology and Medical Oncology, Oncologic Center, Paracelsus Medical University Salzburg, Salzburg 5020, Austria. r.greil@salk.at.|Salzburg Cancer Research Institute with Laboratory of Immunological and Molecular Cancer Research and Center for Clinical Cancer and Immunology Trials, Salzburg 5020, Austria. r.greil@salk.at.|Cancer Cluster Salzburg, Salzburg 5020, Austria. r.greil@salk.at.
authors.forename: Richard
authors.initials: R
authors.lastname: Greil
mesh_terms: D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D005260:Female; D015870:Gene Expression; D020869:Gene Expression Profiling / Q000379:methods / Q000592:standards; D020043:Genes, Essential; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D009362:Neoplasm Metastasis; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods / Q000592:standards
chemical_list: C505362:MIRN16 microRNA, human; C532045:MIRN29a microRNA, human; D035683:MicroRNAs
keywords: breast cancer; control; endogenous control; housekeeper; miR-16; miR-16-5p; microRNA
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