Senologie - Zeitschrift für Mammadiagnostik und -therapie 2015; 12 - A127
DOI: 10.1055/s-0035-1550568

Verification of the breast cancer progression-associated miRNA hsa-miR-199a-5 p using NanoString® platform

S Schultz 1, H Bartsch 2, K Petat-Dutter 2, M Bonin 3, H Seeger 4, S Kahlert 5, U Vogel 6, T Fehm 1, K Soltar 2, D Niederacher 1, H Neubauer 1
  • 1Department of Obstetrics and Gynaecology, Heinrich-Heine-University Duesseldorf, Duesseldorf, Deutschland
  • 2Institute of Pathology, Ludwig-Maximilians-University Munich, Muenchen, Deutschland
  • 3Department of Medical Genetics, Eberhard-Karls-University Tuebingen, Microarray Facility, Tuebingen, Deutschland
  • 4Department of Obstetrics and Gynaecology, Eberhard-Karls-University Tuebingen, Tuebingen, Deutschland
  • 5Department of Obstetrics and Gynaecology, Ludwig-Maximilians-University Munich, Muenchen, Deutschland
  • 6Institute of Pathology, Eberhard-Karls-University Tuebingen, Tuebingen, Deutschland

A fundamental and clinically important step during breast tumourigenesis is the transition from ductal carcinoma in situ (DCIS) to invasive ductal carcinoma (IDC). Improved knowledge of this transition from pre-invasive to invasive breast cancer will pave the way for novel preventative and therapeutic strategies.

We have previously reported on differential expression of the miRNA hsa-miR-199a-5 p in 15 matched pairs of DCIS and IDC areas isolated by laser capture microdissection (LCM) from formalin fixed and paraffin embedded (FFPE) breast cancer tissues using Illumina miRNA BeadChip microarray platform. Differential expression of hsa-miR-199a-5 p was validated by quantitative RT-PCR in additional independent DCIS/IDC sample pairs from 25 breast cancer patients. Knock down of hsa-miR-199a-5 p in invasive MDA-MB-231 and TMX2 – 28 breast cancer cells using a specific inhibitor significantly reduced invasiveness by approx. 73% and 71%, respectively (P < 0.01 and P < 0.05).

Now we report on experiments to validate differential expression of hsa-miR-199a-5 p using a new platform – NanoString® (nCounter® miRNA Expression Assay, NanoString Technologies®). The NanoString® System is an automated, digital detection and counting system which uses a novel barcoding technology to directly profile up to 800 miRNAs simultaneously from a single sample. Total RNA from 6 DCIS/IDC FFPE tumours was used for miRNA expression analysis.

This analysis resulted in 10 differentially expressed miRNAs including hsa-miR-199a-5 p which is upregulated in IDC (P < 0.05). Besides hsa-mir-199a-5 p, hsa-miR-222 is significantly differentially expressed between DCIS and IDC which could be found in both expression data sets (Illumina® and NanoString®).

In this project we identified candidate progression-associated miRNAs which are differentially expressed between DCIS and IDC. Hsa-miR-199a-5 p was validated in an independent sample cohort and its expression was further verified using the new miRNA expression analysis platform NanoString®. Hsa-miR-199a5-p is influencing in vitro cell invasiveness and may therefore be a potential drugable regulator of tumour progression and invasion in breast cancer.