Differential Expression of miR-21, miR-205, miR-221, miR-222, and miR-150 in Molecular Subtypes of Breast Cancer
AR Poongothai, Xu Ling, P Sudipta, Krishan Kalra, Raju K Pillai, Mahul B Amin, Shikha Bose. Biogenex Lab, Freemont, CA; Cedars-Sinai Medical Center, Los Angeles, CA
Background: microRNAs (miRNAs) are short RNA molecules that are involved in many critical cellular processes including oncogenesis. Different cancer types and subtypes at different stages of progression display unique miRNA profiles that may be used as diagnostic, prognostic and therapeutic tools. The detection of miRNA in clinical samples has been difficult, requiring total RNA extracts which lack critical spatial information. In situ hybridization (ISH) allows direct assessment of malignant cells. Evaluation of miRNA profiles in breast cancer (BR CA) holds promise for improving understanding of pathogenesis and therapeutic outcome. This pilot study is designed to assess the feasibility of miRNA profiling in formalin fixed paraffin embedded (FFPE) BR CA samples using in-situ hybridization (ISH) for miR-150, -21, -205, -221 and -222.
Design: 18 samples of FFPE BR CA and 10 normal breast samples were studied. BR CA were molecularly subtyped using immunostains for Estrogen Receptor (InsiteÒER), Progesterone Receptor (InsiteÒPR), and HER-2/neu (Her2)) with SuperSensitiveTM Polymer-HRP IHC Detection System (all BioGenex). Sub categorization resulted in 5 ER/PR positive (+), 5 Her2+, and 8 triple negative (TN) BR CA. Cases were subjected to ISH using FAM-labeled Hsa-miR-1 and Hsa-miR-145 probes (BioGenex) followed by Super Sensitive ISH Detection Kit (BioGenex, DF400-YAX). The signal was amplified with anti-fluorescein antibody and poly-HRP labeled secondary antibody,and visualized with DAB chromogen. Scramble probes served as negative control.
Results: TN BR CA showed up-regulation of miR-21 (2/4 cases), miR-221 (4/8 cases), miR-222 (5/8 cases) and down-regulation of miR-205. miR-150 was up-regulated in Her2+ BR CA but reduced in ER/PR+ BR CA. Normal breast tissue showed low to negative expression of miRNAs.
Figure1: BR CA positive for miR-221 (left panel), negative with scramble probe (right panel)
Conclusions: miRNA profiling is feasible in FFPE samples of BR CA using chromogenic ISH. Although no significant difference was noted in the expression of the various miRNAs in the different molecular subtypes, the presence of differential staining warrants additional studies on larger number of cases to determine prognostic/predictive value.
Wednesday, March 6, 2013 1:00 PM
Poster Session VI # 10, Wednesday Afternoon