MicroRNA profile in very young women with breast cancer.

BACKGROUND: Breast cancer is rarely diagnosed in very young women (35 years old or younger), and it often presents with distinct clinical-pathological features related to a more aggressive phenotype and worse prognosis when diagnosed at this early age. A pending question is whether breast cancer in very young women arises from the deregulation of different underlying mechanisms, something that will make this disease an entity differentiated from breast cancer diagnosed in older patients. METHODS: We performed a comprehensive study of miRNA expression using miRNA Affymetrix2.0 array on paraffin-embedded tumour tissue of 42 breast cancer patients 35 years old or younger, 17 patients between 45 and 65 years old and 29 older than 65 years. Data were statistically analyzed by t-test and a hierarchical clustering via average linkage method was conducted. Results were validated by qRT-PCR. Putative targeted pathways were obtained using DIANA miRPath online software. RESULTS: The results show a differential and unique miRNA expression profile of 121 miRNAs (p-value <0.05), 96 of those with a FDR-value <0.05. Hierarchical clustering grouped the samples according to their age, but not by subtype nor by tumour characteristics. We were able to validate by qRT-PCR differences in the expression of 6 miRNAs: miR-1228*, miR-3196, miR-1275, miR-92b, miR-139 and miR-1207. Moreover, all of the miRNAs maintained the expression trend. The validated miRNAs pointed out pathways related to cell motility, invasion and proliferation. CONCLUSIONS: The study suggests that breast cancer in very young women appears as a distinct molecular signature. To our knowledge, this is the first time that a validated microRNA profile, distinctive to breast cancer in very young women, has been presented. The miRNA signature may be relevant to open an important field of research in order to elucidate the underlying mechanism in this particular disease, which in a more clinical setting, could potentially help to identify therapeutic targets in this particular set of patients.

High stability of microRNAs in tissue samples of compromised quality.

Degradation of tissue samples limits performing RNA-based molecular studies, but little is known about the potential usefulness of samples of compromised quality for studies focused on miRNAs. In this work we analyze a series of cryopreserved tissue samples (n = 14), frozen samples that underwent a severe thawing process (n = 10), and their paired formalin-fixed paraffin-embedded (FFPE) tissue samples (n = 24) from patients with breast cancer obtained during primary surgical resection and collected in 2011. Quality and integrity analyses of the total and small fraction of RNA were carried out. Recovery of specific RNA molecules (miRNAs hsa-miR-21, hsa-miR-125b, and hsa-miR-191; snoRNA RNU6B; and mRNAs GAPDH and HPRT1) was also analyzed by quantitative RT-PCR. Our results suggest that visualisation of the small RNA electrophoretic profiles obtained using the Agilent 2100 bioanalyzer makes it possible to differentiate between the three groups of samples (optimally frozen, thawed, and FFPE). We demonstrate that specific miRNA molecules can be similarly recovered from different tissue sample sources, which supports their high degree of stability. We conclude that miRNAs are robustly detected irrespective of the quality of the tissue sample. In this regard, a word of caution should be raised before degraded samples are discarded: although prior quality assessment of the biological material to be analyzed is recommended, our work demonstrates that degraded tissue samples are also suitable for miRNA studies.