Abstract 2395: Differentiation of carcinoma ex pleomorphic adenoma from pleomorphic adenoma by TERT promoter hypermethylation and elevated TERT mRNA expression

Abstract

Abstract The purpose of this study was to determine whether i) genetic or ii) epigenetic aberrations of the TERT promoter, iii) elevated TERT mRNA level or iv) abnormal p53 expression pattern effectively distinguishes cases of carcinoma ex pleomorphic adenoma (CA ex PA) from pleomorphic adenoma (PA). PA is the most common salivary gland tumor which in rare cases progresses into an epithelial malignancy, termed CA ex PA. Fusion genes involving PLAG1, or less commonly HMGA2, are common in both PA and CA ex PA and are thought to be early genetic drivers in their development. Although the key molecular events which occur during malignant transformation of PA are still unknown, some genetic and epigenetic changes proposed to play a role include amplification of PLAG1, HMGA2, MYC, ERBB2, or chromosome 12q and aberrant promoter hypermethylation of RASSF1, especially in combination with promoter hypermethylation of p16, TERT and WT1. The importance of TERT promoter hypermethylation for malignant transformation in PA may be underappreciated since hypermethylation of that region most critical for malignancy in several cancers, termed the upstream of transcriptional start site (UTSS), has not previously been assayed in PA or CA ex PA. Point mutations or genomic rearrangement in the TERT promoter and elevated TERT mRNA levels are also characteristic of many cancers. We used a panel of 10 PA and 6 CA ex PA clinical samples to determine whether these alterations of TERT, or aberrant expression of TERT mRNA or p53 protein, could effectively distinguish CA ex PA from PA. UTSS hypermethylation in each sample was determined by treatment of genomic DNA with sodium bisulfite followed by MassARRAY and Sanger sequencing. Mutation and arrangement of TERT were screened for by Sanger sequencing and fluorescence in situ hybridization, respectively. p53 expression pattern was visualized by immunohistochemistry and TERT mRNA levels were measured by RT-qPCR. Mutation or rearrangements of the TERT promoter were not observed. However, all 6 CA ex PA samples harbored hypermethylated TERT promoter and had much higher TERT mRNA levels than the PA samples. Two of these samples also had aberrant p53 expression patterns, suggesting the presence of a TP53 missense mutation. None of the PA samples had aberrant p53 expression patterns, although 2 had hypermethylated TERT promoter and elevated TERT mRNA levels as compared with the other PA samples. The significance of this for long term outcome in these patients was unclear. Overall, these results suggest that TERT is commonly upregulated in CA ex PA and that this upregulation is often mediated through hypermethylation of the TERT promoter UTSS. Future large scale studies are warranted to determine the usefulness of TERT promoter hypermethylation for differentiating benign PA from CA ex PA, or the PAs at risk of malignant transformation. Citation Format: Seungjae Lee, Sumit Borah, Kurt Patton, Armita Bahrami. Differentiation of carcinoma ex pleomorphic adenoma from pleomorphic adenoma by TERT promoter hypermethylation and elevated TERT mRNA expression [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 2395. doi:10.1158/1538-7445.AM2017-2395