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

• Published in: Clinica chimica acta 2005-12, Vol.362 (1), p.85-93
• Main Authors: Santonocito, Concetta, Concolino, Paola, Lavieri, Maria Michela, Ameglio, Franco, Gentileschi, Stefano, Capizzi, Rodolfo, Rocchetti, Sandro, Amerio, Pierluigi, Castagnola, Massimo, Zuppi, Cecilia, Capoluongo, Ettore
• Format: Article
• Language: English
• Subjects: Biomarkers, Tumor - genetics; Biomarkers, Tumor - metabolism; Female; Humans; L-Lactate Dehydrogenase - metabolism; Male; Melanoma; Melanoma - blood; Melanoma - genetics; Melanoma - metabolism; Melanoma - pathology; Monophenol Monooxygenase - genetics; Neoplasm Staging; Nerve Growth Factors - genetics; Nerve Growth Factors - metabolism; Nested-PCR; Phosphopyruvate Hydratase - genetics; Phosphopyruvate Hydratase - metabolism; Real-time; Reverse Transcriptase Polymerase Chain Reaction - methods; RNA, Messenger - blood; RNA, Messenger - genetics; S100 Calcium Binding Protein beta Subunit; S100 Proteins - genetics; S100 Proteins - metabolism; Sensitivity and Specificity; Tyrosinase mRNA
• ISSN: 0009-8981; 1873-3492
• DOI: 10.1016/j.cccn.2005.05.031

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. Few commercial methods are available for the detection of tyrosinase mRNA in blood. Comparison between two RT-PCR-nested methods with a third one based on real-time methodology, for detection and quantitation of tyrosinase transcripts, respectively. 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. 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. Tyrosinase mRNA detection may be therefore employed to monitor the melanoma patients over time in function of response to therapy.