Merkel Cell Carcinoma Expression Profiling Demonstrates No Significant Difference between MCPyV Positive and Negative Tumors.
Eric J Duncavage, John D Pfeifer. University of Utah, Salt Lake City; Washington University, St. Louis, MO
Background: Merkel Cell Carcinoma (MCC) is a rare and often fatal cutaneous malignancy that has been shown to harbor clonally-integrated Merkel Cell Polyomavirus (MCPyV) in up to 80% of cases. While morphologically identical, the molecular differences between MCPyV positive and negative MCCs are unknown. We used gene expression profiling to examine the differences between MCPyV positive and negative MCC and to compare the MCC expression profile to published data sets.
Design: Genomic DNA and total RNA were first extracted from formalin-fixed, paraffin-embedded tissue blocks. MCC cases were tested for the presence of MCPyV by PCR using the previously published MCVPS1 primer set. Cases that were MCPyV-positive were further tested by PCR viral-deletion mapping to confirm the presence of truncating large T antigen deletions. Next, a set of 10 MCC cases was selected, consisting of 6 MCPyV-positive and 4 MCPyV-negative MCCs. RNA from these cases was then amplified using the NuGen Ovation FFPE kit and run on Affymetrix Human Exome 1.0 ST arrays. The resulting expression data were clustered using Spotfire and the R statistical package. Confirmation of target genes was performed by TaqMan RT-PCR with 500ng of cDNA on a larger set of 16 MCPyV-positive and 12 negative MCCs.
Results: While there was little overall difference in expression between MCPyV positive and negative MCC, supervised hierarchical clustering defined a cluster of 7 genes with significant differences in expression including NF2, TSPAN15, CMYA5, CCDC55, NAPG, and two ESTs. However, TaqMan PCR for NF2, TSPAN15, CMYA5, and CCDC55 showed no significant difference between MCPyV positive and negative MCC in a larger set of 28 MCC cases. Non-supervised clustering of all MCC cases with published expression data including 158 normal tissues samples revealed an expression pattern most similar to mesoderm-derived tissues and not ectoderm-derived tissues.
Conclusions: Using array-based gene expression profiling from FFPE tissue, we could not detect a reproducible significant difference between MCPyV-positive and negative MCC in the 28 cases tested. This finding is consistent with the so-called 'hit and run' hypothesis in which MCPyV induces host DNA damage early in the pathogenesis of MCC but is not required for disease progression. In addition, the finding that MCC RNA expression profiles are most similar to mesodermal-derived tissue casts some doubt on the currently accepted histogenesis of MCC.
Wednesday, March 2, 2011 1:00 PM
Poster Session VI # 97, Wednesday Afternoon