Comparison between three molecular methods for detection of blood melanoma tyrosinase mRNA. Correlation with melanoma stages and S100B, LDH, NSE biochemical markers

Background: The molecular monitoring of circulating tumor cells by reverse transcriptase-PCR (RT-PCR) for patients with melanoma, is still under debate. It may be affected by: a) pre-analytical variability, b) frequency of melanoma-associated gene transcripts and c) the reliability of the methods employed. Pew commercial methods are available for the detection of tyrosinase mRNA in blood. Objective: Comparison between two RT-PCR-nested methods with a third one basest on real-time methodology, for detection and quantitation of tyrosinase transcripts, respectively. Methods: Sixty-two melanoma patients with different AJCC stages and 20 healthy subjects were enrolled. All blood samples were extracted in duplicate with two different methods. Two nested-PCR methods (one commercial and one in house) plus a real time commercial kit were employed. Results: The two nested PCR methods employed were overimposable, specific and sensitive, at least in the stage III, where there was a concordance between sentinel lymph nodes detection and blood tyrosinase positivity. The different extraction methods did not affect the quality of results, while the commercial real-time kit cannot be used. Conclusion: Tyrosinase mRNA detection may be therefore employed to monitor the melanoma patients over time in function of response to therapy. (c) 2005 Elsevier B.V. All rights reserved.