Circulating tyrosinase mRNA detection in a preclinical xenograft mouse melanoma model and in advanced melanoma patients

M. R. GABRI; V. VAZQUEZ; L. L. OTERO; M. A. CASTRO; G. CINAT; D. E. GOMEZ; D. F. ALONSO

Chicago, Illinois

Congreso; 44th American Society for Clinical Oncology Annual Meeting; 2008

American Society for Clinical Oncology

Background. Although it is still under debate, molecular detection of circulating tyrosinase is believed to be a good prognostic marker for melanoma progression. Methods. We have designed high-affinity primers to amplify the human tyrosinase mRNA sequence to test the value of molecular detection of circulating melanoma cells by reverse transcription-PCR. The optimization process included in vitro settings and in vivo studies in a xenograft mouse model. Results. We were able to detect tyrosinase expression with as few as 40 pg and 1.5 pg of total RNA from SKmel human melanoma cells, using PCR and nested PCR, respectively. Human tyrosinase expression was found in the blood of nude mice bearing s.c. SKmel tumors, and expression bands were stronger after manipulation of the tumor mass. In this melanoma model, Tyrosinase expression declined in blood 6 hours after a direct intravenous injection of SKmel cells. A preliminary study in human blood samples demonstrated a baseline positive tyrosinase determination in 64% (16/25) of advanced melanoma patients using the RT-PCR nested assay. Baseline tyrosinase expression was significantly associated with disease progression after 12 months, and sequential determination during follow up of the remaining disease-free patients showed a progressive increase of negative results. Conclusions. We have developed sequential in vitro and in vivo preclinical validations to improve the optimization of the detection system. Our data indicates that molecular detection of circulating tyrosinase mRNA in advanced melanoma patients could be considered a prognostic parameter.