Preclinical Studies on Detection of Circulating Melanoma Cells in Patients: Telomerase as a Recognition Marker of Malignancy

Abstract

Preclinical studies based on a “simulation design”, were performed with cultured melanoma cells prelabeled with ⁵¹Cr, added to normal blood and subjected to separation and recognition steps. Mononuclear cells (MNC) were isolated on ficollhypaque gradient, and melanoma cells were separated from lymphocytes using anti- CD45 immunomagnetic beads. Malignant cells were then recognized by measuring telomerase activity (TRAP and TRAP-ELISA assays). It was found that: (a) recovery of prelabeled cells present in MNC did not exceed 75%; (b) further recovery of prelabeled cells after separation from lymphocytes did not exceed 68%. Therefore, the overall recovery of prelabeled cells did not exceed 48%; (c) the entire procedure was able to reliably detect as few as 30 malignant cells added to normal blood, providing a telomerase signal significantly higher than that found in absence of melanoma cells. These results furnish the technical bases for developing a tumor detection assay in the blood of melanoma patients.

Keywords:

• Melanoma
• circulating cells
• telomerase
• immunomagnetic beads
• CD45
• TRAP