Abstract Somatic mutation of the promoter region of the telomerase reverse transcriptase (TERT) gene was recently reported as a potential mechanism by which telomerase becomes activated in cancer cells. Here, we aimed to assess the frequency and spectrum of TERT promoter mutations in bladder cancer and detect mutations in voided urine. A SNaPshot assay designed to simultaneously interrogate two positions (-124bp and -146bp) previously reported as mutated in melanoma was used to screen 47 bladder cancer cell lines and 262 tumors. Forty cell lines and 201 tumors carried mutations at either -124bp or -146bp. Mutations at these positions were mutually exclusive. A germline mutation at -57bp (T>G) was previously reported in a melanoma-prone family. A second SNaPshot assay was designed to interrogate position -57bp and we screened all samples that were wildtype at positions -124bp and -146bp. Mutations at -57bp were detected in two cell lines and eight tumors. Matched blood samples were available for six of the tumors and one cell line, and somatic mutation status was confirmed. Sanger sequencing of the TERT core promoter in samples that were wildtype at positions -57bp, -124bp and -146bp revealed a further nine somatic mutations. Four mutations at positions -45bp, -72/-77bp, -141bp, and -54bp were novel, and all but one (-54bp) created new binding sites for ETS/TCF transcription factors. Overall, TERT promoter mutations were detected in 89% of cell lines and 83% of tumors. There was no association of TERT promoter mutation status with stage (p=0.1089, chi-square test) or grade (p=0.2438, chi-square test). As mutations are highly frequent in both non-muscle-invasive and invasive bladder cancer, these represent potential biomarkers for urine-based disease monitoring. For sixty-four tumors used in this study, matched urine samples were also collected prior to surgery. Fifty-three of the tumors had mutations at -124bp or -146bp and using the -124bp/-146bp SNaPshot assay we were able to detect 51 of these in the corresponding urine samples including all mutations (n=23) in 29 patients with low-grade noninvasive tumors. The use of TERT promoter mutation analysis in combination with other DNA-based biomarkers could markedly improve the sensitivity for urine-based detection of recurrent disease. Citation Format: Carolyn D. Hurst, Fiona M. Platt, Margaret A. Knowles. TERT promoter mutations are highly prevalent in bladder cancer and represent a potential new urinary biomarker. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2240. doi:10.1158/1538-7445.AM2014-2240
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