Direct visualization of the clonal progression of primary cutaneous melanoma: application of tissue microdissection and comparative genomic hybridization.

Human cutaneous malignant melanoma progresses through a series of well defined clinical and histopathological stages. It has been assumed that the neoplastic progression of this disease advances from a common acquired nevus or dysplastic nevus through the primary radial growth phase (RGP), primary vertical growth phase (VGP), and finally to distant metastasis. However, it has never been directly shown that VGP is clonally derived from RGP. Furthermore, it has not been possible previously to conduct a detailed genetic analysis on pure tumor cells from archival material because the lesions are a heterogeneous mixture of normal and neoplastic cells, and the entire specimen must be excised and fixed for clinical diagnosis. This report describes a new approach designed to identify DNA copy number changes in tumor cells from a series of progressive primary stages of cutaneous melanoma archival biopsies. Under direct high-power visualization, cells are procured with a sterile needle from highly specific areas of the tissue section. DNA is extracted from microdissected cells (normal, RGP, and VGP), PCR amplified, fluorescently labeled, and examined by comparative genomic hybridization to determine DNA copy number changes. Data obtained from three representative cases suggest a clonal derivation of VGP cells from RGP. This approach could be useful in identifying the sequence of genetic changes in progressive cutaneous melanoma stages.