miR-205 Is Downregulated in Cutaneous Malignant Melanoma
MS Abi Daoud, J Chen, N Zhang, H Feilotter, V Tron. Queen's University, Kingston, ON, Canada
Background: The molecular mechanisms regulating melanoma progression are not well understood. Deregulation of microRNA (miRNA) expression has been observed in many diseases, including cancer. We speculated that miRNAs are deregulated in malignant melanoma, and specific miRNAs are involved in melanoma tumorigenesis and progression.
Design: To address this, we began by comparing the miRNA expression profiles of benign and malignant human melanocytic tumors, including nevi (n=8), primary melanoma (n=8), and metastatic melanoma (n=9) using archival formalin fixed, paraffin embedded (FFPE) tissue. We have previously confirmed the validity of using FFPE tissue to profile miRNAs using an array platform. The Agilent miRNA microarray platform was used to generate miRNA expression profiles, and Significance Analysis of Microarray (SAM) was applied to identify differentially expressed miRNA. Unsupervised hierarchical clustering was able to separate nevi and melanoma samples. We found both up-regulated and down-regulated miRNAs in melanoma. One of the most significantly down regulated miRNAs in melanoma was miR-205, showing a 0.06 fold change in metastatic melanoma, and a 0.23 fold change in primary melanoma, when compared to the expression in benign nevi. TaqMan qRT-PCR analysis validated the down regulation of miR-205 in melanoma (delta Ct 10.53 1.68) over benign nevus (delta Ct 3.98 0.23), p 0.007. To investigate direct downstream targets of miR-205, we performed an in-silico analysis using TargetScan 4.2 software. Two potentially relevant targets to miR-205 were identified, Bcl-2 and c-Kit. A TMA was constructed using the same tumors in which miRNA expression was performed, and immunohistochemistry staining for Bcl-2 and c-Kit conducted. We then ran a Spearman analysis, attempting to show the expected negative correlation between miR-205 array expression levels and protein expression.
Results: A negative correlation was noted between c-Kit expression and miR-205 levels in metastatic melanoma, and not nevi or primary melanoma, but it did not reach significance (-0.268, NS). However, a significant negative correlation was found between miR-205 and Bcl-2 expression in metastatic melanoma samples (-0.818, p0.007), but not in nevi or primary melanoma.
Conclusions: Our studies suggest that miR-205 is deregulated in malignant melanoma in a progression specific manner and that miR-205 loss may be linked to increased Bcl-2 expression.
Monday, March 9, 2009 9:30 AM
Poster Session I Stowell-Orbison/Autopsy Award # 72, Monday Morning