Our evaluation of miRNA expression in archived CRC tissues found similar levels of all six examined miRNAs in tissues stored for over 28 years. An earlier evaluation by Xi et al. (5
) involved analysis of the stability of miR-181b in 40 FFPE CRC samples, and suggested that the expression of miR-181b was stable over a 10 year period. This report, as well as ours, indicates that miRNAs are stable in FFPE tissues stored for long periods of time. Another study by Siebolts et al. (6
) evaluated expression of miR-16 and miR-122a in only three CRC (among 88 different types of malignancies) of frozen and their corresponding FFPE tissues and found no significant differences in the expression levels of these miRNAs. Additionally, there are studies reporting similar yields and stability of miRNAs from matched frozen and FFPE samples (4
). However, the study by Siebolts et al. suggested that specifically the miRNA-16 levels in FFPEs samples of lymph nodes (n=40) stored up to 27 years were decreased nearly by 50%. In contrast, the present investigation is the first to establish the stability of six differentially expressed and potentially clinically relevant miRNAs (miR-20a, miR-21, miR-106a, miR-181b, miR-203, and miR-324-5p) measured by qRT-PCR in a large series of FFPE CRC specimens (n=345) stored for more than 28 years.
Although RNAs in fresh or frozen tissues are relatively stable, there are substantial logistical problems involved in the acquisition of these samples. Conversely, FFPE tissues are a readily available source of samples for retrospective studies to demonstrate correlations between molecular markers and clinical outcomes (7
). Therefore, it would be advantageous to use such tissues for investigations on the clinical value of miRNAs. The lack of sufficient data on the quality of the RNA derived from FFPE tissues due to fixation-related issues, however, is impeding preclinical translational studies of miRNA in cancer. In the fixation process, RNA can form cross-links with protein, and longer RNA molecules are more likely to have cross-links than smaller molecules, such as miRNAs (4
); therefore, miRNAs may not be affected by the fixation. As observed in our study, similar expression levels of the miRNAs in FFPE tissues stored over long periods also support this premise. Currently, we are correlating the miRNA expression profiles with other molecular markers of CRCs and with tumor progression and clinical outcomes. In summary, the findings of the present investigation suggest that miRNAs are stable in FFPE CRC tissues stored for long periods and provide a rationale for using FFPE tissues in investigations involving miRNA profiling.