Abstract 2860: Defining the role of NRAS in melanoma maintenance using a novel inducible mouse model
Authors: VanBrocklin Matthew W. , Robinson James P. , McKinney Andrea J. , Holmen Sheri L. 
Published: June 2012

Abstract
As treatments move into the era of targeted therapy, there is a growing need to understand the alterations that result in melanoma formation, maintenance, and progression. To this end, we have developed a somatic cell gene transfer model of melanoma based on the RCAS/TVA system that allows for tissue- and cell-specific targeted infection of mammalian cells through ectopic expression of the viral receptor. This system utilizes a retroviral vector, RCASBP(A), derived from the avian leukosis virus (ALV). The receptor for RCASBP(A) is encoded by the tv-a gene and is normally expressed in avian cells; infection results in integration of the virus into the genome of dividing cells and expression of delivered genes. Since the virus is replication-defective in mammalian cells, these cells can be infected with multiple ALV-derived viruses. This allows efficient modeling of melanoma because multiple oncogenic alterations can be introduced into the same cell. The dopachrome tautomerase (DCT) promoter was chosen to drive expression of the viral receptor TVA since this gene is expressed early in melanocyte development when the cells are mitotically active. A significant percentage of both familial and sporadic melanomas have mutations that functionally inactivate both INK4a and ARF; therefore, DCT-TVA mice were crossed to Ink4a/Arflox/lox mice. Activated NRAS oncogenes have also been detected in approximately 20% of human melanomas. Therefore, as a proof-of-principle, newborn mice were injected subcutaneously with retroviruses containing Cre-recombinase and NRASQ61R. Whereas no tumors were detected in TVA-negative mice, melanomas were visible in DCT-TVA;Ink4a/Arflox/lox mice as early as three weeks of age. Within three months, more than two-thirds of DCT-TVA;Ink4a/Arflox/lox mice developed melanoma that was histologically similar to the human disease. To allow for regulation of NRAS expression post-delivery using the tetracycline (tet)-regulated system, a tet-responsive element (TRE) was inserted upstream of the NRAS gene in the viral vector. Expression from the TRE requires the presence of a tetracycline transcriptional activator (tTA) such as Tet-off or a reverse tTA (rtTA) such as Tet-on. In the context of Tet-off, the Tet-responsive gene is repressed in the presence of Dox. Doxycycline (Dox) was administered to suppress NRAS expression once tumors reached 1,000 mm3. Within four days, Dox induced substantial regression of NRAS and Cre induced melanomas in DCT-TVA/Ink4a/Arf lox/lox mice. Tumors regressed in all mice treated with Dox. Interestingly, the tumors did not disappear completely but regressed to ∼5 mm3 and became dormant. This model will be used to study tumor dormancy, define mechanisms of resistance, and test novel combination therapies. Our long-term goals are to translate the knowledge gained from these studies into improvements in molecular targeted therapies for the treatment of advanced melanoma.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2860. doi:10.1158/1538-7445.AM2011-2860