Background Assays identifying circulating tumor cells (CTC) allow noninvasive and sequential monitoring of the status of primary or metastatic tumors, potentially yielding clinically useful information. However, the effect of radiation therapy (RT) on CTC in patients with non-small cell lung cancer (NSCLC) to our knowledge has not been previously explored. Methods We describe here results of a pilot study of 30 NSCLC patients undergoing RT, from whom peripheral blood samples were assayed for CTC via an assay that identifies live cells, via an adenoviral probe that detects the elevated telomerase activity present in almost all cancer cells but not normal cells, and with validity of the assay confirmed with secondary tumor-specific markers. Patients were assayed prior to initiation of radiation (Pre-RT), during the RT treatment course, and/or after completion of radiation (Post-RT). Results The assay successfully detected CTC in the majority of patients, including 65% of patients prior to start of RT, and in patients with both EGFR wild type and mutation-positive tumors. Median counts in patients Pre-RT were 9.1 CTC/mL (range: undetectable - 571), significantly higher than the average Post-RT count of 0.6 CTC/mL (range: undetectable - 1.8) (p < 0.001). Sequential CTC counts were available in a subset of patients and demonstrated decreases after RT, except for a patient who subsequently developed distant failure. Conclusions These pilot data suggest that CTC counts appear to reflect response to RT for patients with localized NSCLC. Based on these promising results, we have launched a more comprehensive and detailed clinical trial.

No abstract

BackgroundCirculating tumor cell (CTC) detection and genetic analysis may complement currently available disease assessments in patients with melanoma to improve risk stratification and monitoring. We therefore sought to establish the feasibility of a telomerase-based assay for detecting and isolating live melanoma CTCs.MethodsThe telomerase-based CTC assay utilizes an adenoviral vector that, in the presence of elevated human telomerase activity, drives the amplification of green fluorescent protein. Tumor cells are then identified via an image processing system. The protocol was tested on melanoma cells in culture or spiked into control blood, and on samples from patients with metastatic melanoma. Genetic analysis of the isolated melanoma CTCs was then performed for BRAF mutation status.ResultsThe adenoviral vector was effective for all melanoma cell lines tested with sensitivity of 88.7% (95%CI 85.6-90.4%) and specificity of 99.9% (95%CI 99.8-99.9%). In a pilot trial of patients with metastatic disease, CTCs were identified in 9 of 10 patients, with a mean of 6.0 CTCs/mL. At a cutoff of 1.1 CTCs/mL, the telomerase-based assay exhibits test performance of 90.0% sensitivity and 91.7% specificity. BRAF mutation analysis of melanoma cells isolated from culture or spiked control blood, or from pilot patient samples was found to match the known BRAF mutation status of the cell lines and primary tumors.ConclusionsTo our knowledge, this is the first report of a telomerase-based assay effective for detecting and isolating live melanoma CTCs. These promising findings support further studies, including towards integrating into the management of patients with melanoma receiving multimodality therapy.

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

No abstract