A polymerase chain reaction-based semiquantitative assessment of malignant melanoma cells in peripheral blood

Abstract

Malignant melanoma cells can be detected with high sensitivity in peripheral blood of patients using reverse transcription-PCR. The detection of tyrosinase mRNA that is actively expressed only in melanocytes and melanoma cells indicates the presence of melanoma cells in peripheral blood. As shown previously, tyrosinase transcripts can be found in a variety of patients with metastatic malignant melanoma. For semiquantitative analysis of these cells in peripheral blood and evaluation of possible influence of immunotherapy on the amount of circulating cells, we describe an assay combining reverse transcription-PCR and Southern blotting. In this system, the amount of circulating tumor cells was determined by interpolating the amplified tyrosinase signal strength of patient samples to an equivalent tyrosinase signal of diluted SK-mel 28 cells. We found that the amount of circulating tumor cells correlates with the tumor burden. Furthermore, in patients with regression of melanoma metastases after immunotherapy, a decrease of the amount of tumor cells in the peripheral blood was observed. Quantitative estimates of residual disease may be an accurate and sensitive predictor for the clinical course.