Poster presentation, P143

Official XXIst International Pigment Cell Conference website - 21-24 Sept 2011, Bordeaux - France | updated: September 04 2011

Detection, Quantification and Characterisation of PAX3 across the Spectrum from Melanocytes to Melanoma and in Circulating Melanoma Cells Relative to Disease Stage

BACKGROUND AND OBJECTIVES: The aim of this research was to assess PAX3 expression across the spectrum from melanocyte to melanoma and determine its role in tumour progression. The transcription factor PAX3 regulates melanocyte specification from neural crest cells during development but expression in differentiated melanocytes has not been detailed to date. In this study we analysed PAX3 expression in normal skin melanocytes, in melanoma cells of primary and metastatic tissue and in circulating melanoma cells. We then quantified and characterised circulating melanoma cells using a variety of markers including PAX3. METHODS: PAX3 expression was assessed in normal skin melanocytes and melanoma cells, using immunofluorescence and qRT-PCR. qRT-PCR was also used to quantify PAX3 levels relative to other melanoma markers, in patient blood of 300 melanoma patients and 100 healthy volunteers. Frequency and level of expression of melanoma markers in patient blood were correlated with Breslow thickness of the primary tumour and to presence of metastases and results were statistically analysed. Immunomagnetic bead capture and flow cytometry were used to isolate, quantify and characterise circulating cells from patient blood samples. RESULTS: Here we demonstrate PAX3 in normal skin melanocytes as well as in melanoma cells of primary melanoma and metastatic melanoma, and in circulating melanoma cells. Furthermore, circulating melanoma cells were identified in 79% of patients with stage III and IV disease whilst these markers were observed in only 20-30% of early stage patients. Surprisingly, melanoma cells are found in peripheral blood of patients with no clinical evidence of metastatic disease, including patients with early stage disease and patients from whom tumours were removed several years previously. Flow cytometry and immunomagnetic bead capture confirmed the presence and quantity of circulating melanoma cells relative to disease stage. CONCLUSION: PAX3 is a useful marker of melanocytes and melanoma cells, including circulating melanoma cells. As such, PAX3 can be utilised as one of several melanoma markers for quantification and characterisation of circulating melanoma cells and may be a valuable clinical indicator of disease progression and treatment efficacy. This research is funded by grants from the NHMRC of Australia and the Cancer and Palliative Care Research and Evaluation Unit.

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