ORIGINAL ARTICLES: Basic researchMetastasis in an orthotopic murine model of melanoma is independent of RAS/RAF mutationRozenberg, Gabriela I.a b; Monahan, Kimberly B.a; Torrice, Chada b; Bear, James E.c; Sharpless, Norman E.a bAuthor Information Departments of aGenetics bMedicine cHHMI, Cell and Developmental Biology, The Lineberger Comprehensive Cancer Center, University of North Carolina School of Medicine, Chapel Hill, North Carolina, USA Correspondence to Norman E. Sharpless, MD, Departments of Medicine and Genetics, The Lineberger Cancer Center, CB#7295, The University of North Carolina School of Medicine, Chapel Hill, NC 27599-7295, USA Tel: +1 919 966 1185; fax: +1 919 966 8212; e-mail: [email protected] Received 2 September 2009 Accepted 18 December 2009 Supplemental digital content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal's Website (www.melanoma.com). Melanoma Research: October 2010 - Volume 20 - Issue 5 - p 361-371 doi: 10.1097/CMR.0b013e328336ee17 Buy SDC Metrics Abstract Melanoma is the most lethal skin tumor in large part because of a propensity for early metastasis. Good models of this most clinically relevant feature of melanoma are lacking. Here, we report the development of an in-vivo model of metastasis that relies on orthotopic injection of green fluorescent protein-tagged lines in immunodeficient mice, serial intravital imaging of tumor progression, and quantification of distant spread by two-photon laser scanning microscopy, immunohistochemistry, and real-time PCR analysis. Using this system, we report an assessment of the in-vivo growth and metastatic properties of 11 well-characterized human melanoma cell lines. A subset of lines showed rapid in-vivo growth with invasion of host vasculature and distant seeding of viscera in this system. The ability to form metastasis in vivo did not correlate with three-dimensional collagen invasion in vitro. Surprisingly, similar lines in terms of molecular genetic events differed markedly in their propensity to metastasize to distant organs such as brain and lung. In particular, two lines harboring B-RAF mutation and high levels of phosphorylated ERK and AKT were reproducibly unable to form tumors after orthotopic injection. Similarly, two previously identified RAS/RAF wildtype ‘epithelial like’ lines that do not have elevated phosphorylated ERK and AKT or express TWIST1 mRNA still showed a pronounced ability for orthotopic growth and metastatic spread. All the metastatic cell lines in this model showed increased NEDD9 expression, but NEDD9 lentiviral overexpression did not convey a metastatic phenotype on nonmetastatic cells. These data suggest that melanoma metastasis is a molecularly heterogeneous process that may not require epithelial-to-mesenchymal transition or ERK activation, although both may facilitate the process. © 2010 Lippincott Williams & Wilkins, Inc.