BACKGROUND: The treatment of melanoma is rapidly evolving, with many recently introduced systemic therapies. The management of individual patients, however, remains challenging. Decisions on when to initiate and discontinue specific treatments rely on imprecise methods such as imaging studies, and patient outcomes often remain less than optimal. An assay that can be performed conveniently, safely, and serially could usefully complement current assessments of disease burden. Circulating tumor cell (CTC) analysis fulfills all these criteria and may assist in determining true disease status and risk stratification. However, commercial CTC assays that target epithelial cell markers are ineffective for melanoma. We, therefore, investigated the use of a novel telomerase-based assay for melanoma CTCs.
METHODS: The assay utilizes an adenoviral vector that, in the presence of elevated human telomerase (hTERT) activity, drives the expression of green fluorescent protein (GFP). This confers sensitivity and specificity, as telomerase activity is elevated in almost all cancer cells but not elevated in almost all normal cells. The tumor cells are identified via an image processing system that applies cutoff criteria of fluorescent intensity and size. The cells are co-stained with anti-Melan-A to confirm their melanoma origin. The assay was tested on melanoma cells in culture, in control blood, and on samples from 10 patients with metastatic melanoma.
RESULTS: The assay was effective for all melanoma cell lines tested with sensitivity of 92% (95% confidence interval (CI) 84.4-99.1%) and specificity of 99% (95%CI 99.5-99.6%). It was also effective in control blood from healthy volunteers (limit of detection = 1.1 CTCs/mL). In a pilot trial of patients with metastatic disease, CTCs were identified in 9 of 10 patients (the tenth patient was without evidence of disease after treatment) with mean count of 6.0 CTCs/mL (range 0.7-27.1 CTCs/mL). In a linear regression model, CTC counts were significantly increased in patients with greater burden of disease (p = 0.02) and significantly decreased in patients with current or recent cytotoxic chemotherapy (p = 0.02). Interestingly, prior or active immunotherapy did not significantly affect CTC levels. Patient demographics, primary tumor location, and sites of metastases also did not appear to significantly affect CTC levels.
CONCLUSIONS: To our knowledge, this is the first report of a telomerase-based assay for detecting melanoma cells. This assay was effective in identifying melanoma cells in culture and in the peripheral blood of patients with metastatic melanoma. Greater disease burden appears to be associated with increased CTC levels while cytotoxic chemotherapy appears to be associated with decreased CTC levels. Longer term follow-up and serial assays may validate the ultimate usefulness of this assay in the management of patients with melanoma.
Citation Format: Melody Ju, Gary D. Kao, Charles B. Simone, David Steinmetz, Xiangsheng Xu, Louise Aguarin, Wei Xu, Edmund Bartlett, Stephen M. Hahn, Jay F. Dorsey, Giorgos Karakousis. “Capturing the elusive foe”: A novel telomerase promoter-based approach to detect melanoma circulating tumor cells. [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 1871. doi:10.1158/1538-7445.AM2014-1871