Abstract 5555: Metastatic melanoma associated genes identified using the human tumor metastasis RT2 profiler PCR array
Authors: White Kirsten A. , Flores Kristina , Berwick Marianne 
Published: August 2012

Metastatic melanoma is a complicated disease with growing rates of diagnosis and few clinical treatment options currently available. While melanoma results in only 5% of skin cancers, it is the cause of 75% of all skin cancer deaths. In 2008, the American Cancer Society (ACS) reported 62,480 new cases of melanoma. Of these individuals, 8,420 died, including 5,400 men and 3,020 women. According to the ACS for 2010, the 5-year survival rate for stage IV melanoma is 15% to 20% and the 10-year survival is 10% to 15%. To improve survival it is important to establish which genes are involved in the regulation of metastasis in melanoma as potential targets for prevention and/or treatment. Our laboratory compared the differences in the relative expression levels of metastasis genes from two human melanoma cell lines and a normal melanocyte cell line. The normal melanocyte cell line was assayed in duplicate and averaged as an inner plate control in order to validate the assay. Established melanoma, SK-mel-103 and SK-mel-147, were investigated. Template cDNA was prepared from total RNA from these lines using the Qiagen RNeasy Mini kit. The melanoma cell lines were both male and matched on GPR-30 and ER beta expression and B-RAF and N-RAS mutation status but they differed in G2 checkpoint and p16 mutation status. RT Array technology was utilized to compare the relative levels of mRNA from metastasis related genes expressed in SK-mel lines. The Human Tumor Metastasis RT2 Profiler™ PCR Array contains 84 primer pairs that amplify genes involved in human metastasis. Genes on this array include those that regulate cell adhesion, the extracellular matrix components, cell cycle, cell growth and proliferation, apoptosis, and transcription factors and regulators. The PCR array is a set of optimized real-time PCR primer assays on a 384-well plate for pathway focused genes and includes appropriate RNA quality controls. The quantitative PCR was run on the ABI 7900 HT using the real-time PCR cycling program. Total RNA from normal and melanoma tumors were characterized and the relative expression levels for each gene in the two melanoma samples were plotted against each other and against the normal cell line in scatter plots. Gene expression with controls samples was highly correlated. Genes involved in metastasis were up or down regulated in the melanoma cell lines compared to the normal cell lines. Upregulation of genes associated with DNA replication has previously been correlated with metastatic aggressiveness in melanoma cell lines. Conversely the down regulation of genes has been associated with an increase in cellular adhesion and melanocyte differentiation. This research identifies metastatic genes that contribute to the metastatic aggressiveness in the SK-mel metastatic melanoma cell lines. In the future we plan to examine the genes identified in this study for novel drug repurposing experiments.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5555. doi:1538-7445.AM2012-5555