Abstract
Abstract Background: The diagnosis of leptomeningeal metastases is usually based on the finding of malignant cells by cytological examination in the cerebrospinal fluid (CSF). However, cytology is only moderately sensitive, and sensitive and specific cancer biomarkers may improve the detection of tumor cells in the CSF. Expression of telomerase hTERT subunit might be such a marker but its use is hampered by proliferating normal lymphocyte cells. hTERT methylation, characterizing most cancer cells, might be an alternative as proliferating lymphocytes have unmethylated hTERT promoter. The aim of this study was to develop a specific, sensitive, quantitative, and fast method for detection of hTERT methylation and to explore its use as a cancer biomarker in the diagnosis of metastatic tumors in CSF. Materials and methods: hTERT methylation levels were assayed by several quantitative techniques including methylation-sensitive dot blot assay (MS-DBA), methylation-sensitive high resolution melting (MS-HRM), and a newly developed real-time MS-HRM assay. We used MS-HRM assays for the analysis of 59 CSF specimens from 50 patients including 31 CSFs from 23 patients with a known malignancy suspected for leptomeningeal metastasis. Results: MS-HRM assays both allowed hTERT methylation quantification in CSF samples. The MS-HRM detected samples containing more than 10% of hTERT methylated alleles whereas real-time MS-HRM detected methylation below 10%. PCR products were obtained from 54 CSF samples (92%). hTERT methylation was only detected in the CSF from patients with a known malignancy. Conclusion: The real-time MS-HRM analysis is a fast, sensitive, and specific technique for methylation assessment in many diagnostic and research applications. As an adjunct to the traditional examination of CSF, the hTERT MS-HRM approach could be perform as routine diagnosis of leptomeningeal metastases and could become a valuable tool for further management and planning of treatment. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4931.
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