Isolation of invasion-associated cDNAs in melanoma.

Abstract

Metastasis and invasion are key steps in the systemic spread of tumor cells. To identify the genes involved in this process we recently selected highly invasive and weakly invasive cell clones from a melanoma cell line. Both cell clones showed a stable phenotype over more than 40 passages and previous analyses revealed a fivefold difference in their invasive potential in vitro and in tumorigenesis in vivo. To compare gene expression of the two cell clones a cDNA array system (Clontech human cancer cDNA array) was used. Exact quantification of differentially expressed genes in each cell clone was performed by real-time RT-PCR. An evaluation of the array data revealed a total of 36 genes that were more than 1.5-fold differentially expressed, and 26 (72%) of these showed a differential expression pattern by quantitative RT-PCR. Previously known differences in expression patterns, including loss of p16 and HLA I, or equal expression of p73, and RAR alpha, beta and gamma were confirmed by the array data. In addition, reduced expression levels of several cytoskeletal proteins, such as vimentin, gamma-actin, desmin and cytokeratins, in the highly invasive cell clone were reproducibly identified. Other genes strongly upregulated in the highly invasive cell clone included jagged 2, STAT1, tPA and c-myc, whereas MDA-6 (p21), caspase 2 and semaphorin were found to be downregulated. In conclusion, comparative hybridization of cDNA arrays identified a series of novel invasion-associated changes in gene expression and confirmed previously known expression patterns.