Stage-dependent differential expression of microRNAs in colorectal cancer: potential role as markers of metastatic disease.

MicroRNAs (miRs) are short non-coding RNAs that bind complementary sequences in mRNA resulting in translation repression and/or mRNA degradation. We investigated expression of the reported metastasis-associated miRs-335, 206, 135a, 146a, 146b, 10b, 21, let7a and let7b in normal mucosa, non-metastatic and metastatic colorectal cancer (CRC). Expression of target miRs in micro-dissected paraffin embedded tissues was evaluated in 15 primary tumours with adjacent normal tissue from patients that were disease-free at 4 years (cohort A) and 19 paired primary tumours with corresponding liver metastases (cohort B) by quantitative real-time PCR. Increased expression of miR-21, mir-135a and miR-335 was associated with clinical progression of CRC, while miR-206 demonstrated an opposite trend. The levels of mir-21 did not associate with the expression of PTEN, an important tumour suppressor in CRC and one of many putative targets of miR-21, but interestingly was associated with stage of disease in the PTEN expressing tumours. Surprisingly, let7a, a KRAS-targeting miR, showed elevated expression in metastatic disease compared to normal mucosa or non-metastatic disease, and only in KRAS mutation positive tumors. Finally, a prognostic signature of miR 21,135a, 335, 206 and let-7a for detecting the presence of metastases had a specificity of 87% and sensitivity of 76% for the presence of metastases. In summary, we have shown stage-associated differential expression of five out of nine tested metastasis-associated miRs. We have further found that an analysis of these five miRs expression levels in primary tumors significantly correlates with the presence of metastatic disease, making this a potential clinically useful prognostic tool.