Real-Time PCR Validation of Micro-RNA Array Analysis for Renal Neoplasms
A Liang, L Kelly, J Burger, C Sciulli, R Dhir, W LaFramboise. University of Pittsburgh Medical Center, Pittsburgh, PA
Background: Comprehensive screening of microRNA expression has established that these molecules play a critical role in determining cell fate in development, and contribute to tumorigenesis. Therefore, these molecules hold promise as prospective diagnostic and/or prognostic tumor biomarkers. Use of the biomarkers as clinical probes requires validation of array findings and implementation of a test format conducive to routine clinical screening. In our study, we evaluated renal neoplasm miRNA biomarkers identified by microarray analysis using real time qPCR to validate the array findings and test the sensitivity of these molecules as biomarkers in a targeted molecular assay.
Design: Microarray analsis was performed on RNA extracted from a variety of renal neoplasms (clear cell, chromophobe, and papillary RCC; oncocytoma). MiRNAs found to be significantly different between the neoplasms were then analyzed by real time qPCR; these include miR-886-5p, 221, 222, 597, 204, 10a and 508-5p. These miRs include those with both low and high fluorescence expression by microarray analysis. Internal controls (U6 and mir-25) are used for each miRNA analyzed.
Results: For miRNAs with high expression by microarray analysis (204, 221, 222 and 886-5p), there is high correlation between microarray and PCR results.
For microarray results with low expression (508-5p, 597 and 10a) the correlation is significantly weaker. Although a direct statistical comparison between the two methods cannot be performed, the comparison of relative peak height between tumors in each method shows a correlation between testing methods.
Conclusions: Microarray analysis provides a thorough profile of altered miRNAs in a variety of renal neoplasms. Microarray results with high expression are confirmed as significant using qPCR. Furthermore, for future clinical application, qPCR may prove a rapid and specific test for analysis of select miRNAs.
Category: Genitourinary (including renal tumors)
Wednesday, March 24, 2010 9:30 AM
Poster Session V # 89, Wednesday Morning