Abstract 3532: No REST in Merkel cell carcinoma
Authors: Emil Chteinberg, Christopher Sauer, Lisa D. Schiffelers, Jonathan P. Eben, Dorit Rennspiess, Christopher B. Buck, Veronique J. Winnepenninckx, Anna K. Kurz, Ernst J. Speel, Martin Zenke, Axel zur Hausen
Journal: Cancer Research (2017)

Abstract Introduction Merkel cell carcinoma (MCC) is a highly aggressive skin cancer of the elderly and immunosuppressed patients. More than 80% of MCCs are associated with the recently identified Merkel cell polyomavirus (MCPyV). MCCs reveal a trilinear differentiation characterized by neuroendocrine (chromogranin A and synapthophysin), epithelial (e.g. CK20) and pro/pre B-cell lymphocytic (e.g. Pax5 and TdT) marker expression. The cellular origin of MCCs still remains obscure. Based on the combined expression of TdT, Pax5 and immunoglobulins (Ig), including clonal Ig rearrangements we have recently hypothesized that pre/pro B-cells might constitute the cellular origin of MCC. We aimed to understand the neuroendocrine expression of chromogranin A and synapthophysin which is known to be negatively regulated by the Re-1 silencing transcription factor (REST) in MCC cells. We assessed REST expression in MCPyV-positive and -negative MCCs and MCC cell lines in order to gain insights into the regulation of neuroendocrine differentiation in MCC cells. Methods The expression of REST in MCC was tested by immunohistochemistry (IHC) in 22 formalin fixed and paraffin embedded MCC tissues. In addidtion, MCPyV-positive (MKL-1 and MKL-2) and MCPyV-negative (MCC13 and MCC26) MCC cell lines and the B-ALL cell-line REH as positive control were tested for REST expression. RT-PCR was used to confirm the results of the cell lines on the transcriptional level. In addition, the regulation of the REST expression by methylation was investigated with the demethylating agent 5-aza-2’-deoxycytidin. Further the methylation status of the 3 CpG islands of the REST promoter was analysed with a methyl specific PCR. Results: All MCCs except one were completely devoid of REST expression as tested by IHC. All MCCs were negative for REST but synaptophysin and chromogranin A positive. The MCPyV-positive cell lines MKL-1 and MKL-2 did not express REST but indeed expressed both neuroendocrine markers. The MCPyV-negative cell lines MCC13, MCC26 and the B-ALL cell line were positive for REST expression and negative for these neuroendocrine markers. On RNA level these results were confirmed. The demethylation of the CpG islands of the REST promoter did not lead to an increase of the REST expression in MKL-1 and MKL-2 on the transcriptional or translational level. Initial MSP results did not reveal any methylation of the 3 CpG islands of the REST promoter. Conclusion The lack of REST expression in MCC and in MCPyV-positive MCC cell lines, in combination with REST expression in MCPyV-negative cell lines point to an important role of the MCPyV in the regulation of REST expression in MCC. This is emphasized by the results of 5-aza-2’-deoxycytidin treatment of the MCC cell lines. Our data might provide the basis of neuroendocrine gene expression in MCC which possibly originates from early B cell. Citation Format: Emil Chteinberg, Christopher Sauer, Lisa D. Schiffelers, Jonathan P. Eben, Dorit Rennspiess, Christopher B. Buck, Veronique J. Winnepenninckx, Anna K. Kurz, Ernst J. Speel, Martin Zenke, Axel zur Hausen. No REST in Merkel cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3532. doi:10.1158/1538-7445.AM2017-3532