MicroRNA profiles discriminate among colon cancer metastasis

MicroRNAs are being exploited for diagnosis, prognosis and monitoring of cancer and other diseases. Their high tissue specificity and critical role in oncogenesis provide new biomarkers for the diagnosis and classification of cancer as well as predicting patients' outcomes. MicroRNAs signatures have been identified for many human tumors, including colorectal cancer (CRC). In most cases, metastatic disease is difficult to predict and to prevent with adequate therapies. The aim of our study was to identify a microRNA signature for metastatic CRC that could predict and differentiate metastatic target organ localization. Normal and cancer tissues of three different groups of CRC patients were analyzed. RNA microarray and TaqMan Array analysis were performed on 66 Italian patients with or without lymph nodes and/or liver recurrences. Data obtained with the two assays were analyzed separately and then intersected to identify a primary CRC metastatic signature. Five differentially expressed microRNAs (hsa-miR-21, -103, -93, -31 and -566) were validated by qRT-PCR on a second group of 16 American metastatic patients. In situ hybridization was performed on the 16 American patients as well as on three distinct commercial tissues microarray (TMA) containing normal adjacent colon, the primary adenocarcinoma, normal and metastatic lymph nodes and liver. Hsa-miRNA-21, -93, and -103 upregulation together with hsa-miR-566 downregulation defined the CRC metastatic signature, while in situ hybridization data identified a lymphonodal invasion profile. We provided the first microRNAs signature that could discriminate between colorectal recurrences to lymph nodes and liver and between colorectal liver metastasis and primary hepatic tumor

MicroRNA Profiles Discriminate among Colon Cancer Metastasis

MicroRNAs are being exploited for diagnosis, prognosis and monitoring of cancer and other diseases. Their high tissue specificity and critical role in oncogenesis provide new biomarkers for the diagnosis and classification of cancer as well as predicting patients' outcomes. MicroRNAs signatures have been identified for many human tumors, including colorectal cancer (CRC). In most cases, metastatic disease is difficult to predict and to prevent with adequate therapies. The aim of our study was to identify a microRNA signature for metastatic CRC that could predict and differentiate metastatic target organ localization. Normal and cancer tissues of three different groups of CRC patients were analyzed. RNA microarray and TaqMan Array analysis were performed on 66 Italian patients with or without lymph nodes and/or liver recurrences. Data obtained with the two assays were analyzed separately and then intersected to identify a primary CRC metastatic signature. Five differentially expressed microRNAs (hsa-miR-21, -103, -93, -31 and -566) were validated by qRT-PCR on a second group of 16 American metastatic patients. In situ hybridization was performed on the 16 American patients as well as on three distinct commercial tissues microarray (TMA) containing normal adjacent colon, the primary adenocarcinoma, normal and metastatic lymph nodes and liver. Hsa-miRNA-21, -93, and -103 upregulation together with hsa-miR-566 downregulation defined the CRC metastatic signature, while in situ hybridization data identified a lymphonodal invasion profile. We provided the first microRNAs signature that could discriminate between colorectal recurrences to lymph nodes and liver and between colorectal liver metastasis and primary hepatic tumor

miR-103 In Situ Hybridization of Normal Colon (5X), Primary Tumor (5X), Normal Liver (5X), Liver Metastasis (5X, 10X) and Metastatic Lymph node (5X).

<p>In normal colon the arrow is pointing at the lack of miR-103 expression in epithelial cells. In the primary tumor the arrow is pointing at the increased expression of miR-103 in the invasive adenocarcinoma. The arrow in liver recurrence and the two arrows in the lymph node metastases are showing a dramatic increased expression of miR-103 in the metastatic adenocarcinoma epithelia.</p

Experimental procedure and miRs signatures.

<p>Experimental procedure and miRs signatures.</p

qRT-PCR Box-Plots.

<p>Normal colon tissue (NORM), colon adenocarcinomas (ADENOCA), lymphonodal metastasis (POSLYM) and colon liver (LIVERMET) for miR-21(A), miR-31 (B), miR-93 (C), miR-103 (D) and miR-566 (E). A Mann-Whitney test was applied to compare groups. Groups are shown on the boxplots' x-axis, while the delta Ct Values are represented on the y-axis. For each box, the bar represent the Median, the area the 25^th and 75^th percentile and the whiskers of the graph the largest and smallest values. Each P-value bar correspond to a comparison: for each miR the first lower bar refers to the NORM vs ADENOCA comparison; miR-21 second lower bar P-value corresponds to the NORM vs POS LYMPH comparison, while the first upper bar of both miR-21 and -93 corresponds to the NORM vs LIVER MET comparison.</p

miR-21 In Situ Hybridization of Normal Colon (5X), Primary Tumor (5X) and Liver Metastasis (Different magnifications: 5X, 10X, 20X).

<p>As documented in literature, miR-21 showed a weak signal in normal colon respect to adenocarcinoma (P-value = 0.0011) and it was highly expressed in metastasis. MiR-21 expression was detected only in primary adenocarcinoma and metastatic malignant cells.</p