C67
Cell senescence and melanoma progression: immortalization as a late event
Julia K Soo, Alastair D MacKenzie Ross, David M Kallenberg, Heung Chong, Jade Chow, Barry Newell, Dorothy C Bennett
St George's, University of London, London SW17 0RE, UK
Background
Cell senescence is a permanent proliferative arrest following extensive division, oncogene activation or cellular stresses. Evidence for senescence as a key tumour-suppressor mechanism is particularly strong for melanoma, where the predominant susceptibility locus CDKN2A encodes two senescence effectors, p16(INK4A) and ARF. Benign naevi (moles) are common lesions frequently showing activated BRAFV600E, yet static, apparently through oncogene-induced senescence, though this has been disputed. Metastatic melanoma cells, conversely, generally appear immortal. We previously showed loss of immunoreactive p16 and activation of p53 during melanoma progression, yet with little expression of p21 (effector of p53-mediated senescence).
Method
Primary micro-cultures have been sampled from excised pigmented lesions, to confirm directly which are senescent and when in progression melanoma cells become immortal. The culture method uses “feeder” keratinocytes and growth factors. Lesional cells attach and form healthy cultures. Senescence or proliferation are ascertained using growth curves, microscopy and markers of senescence (p16, acidic β-galactosidase and heterochromatic foci containing trimethyl-histone H3-K9).
Results
100% of benign and dysplastic naevus cultures senesced, showing arrested growth and 3/3 senescence markers, though most divided initially. Many thin and thick primary melanoma cultures also showed proliferation then growth-arrest, but only a minority of cells displayed senescence markers. The presence of highly abnormal cell morphology, multiple nuclei, chromosome rearrangements and floating dead cells suggested that telomeric crisis may have been reached. Only a few melanomas appeared immortal.
Conclusion
Some propose that all cancer cells are immortal, but these findings suggest that immortalization is a late event in melanoma progression.
