Intravital Imaging Of Endogenous Brafv600e Melanoma Reveals Plasticity Of Tumor Response And Resistance To Trametinib Over Time

Although the number of therapies entering clinical trials continues to increase, the success rate for targeted therapy studies is quite low due, in part, to a lack of relevant preclinical models. The use of targeted therapy has become a popular method for cancer treatment against mutant BRAF melanoma, as studies show that clinical use of selective inhibitors against MEK and BRAF extend life expectancy. However, melanoma becomes refractory to treatment. Since the tumor and stromal factors that contribute to drug resistance remain poorly understood, our ability to treat patients with advanced disease remains limited. Current genetically engineered murine (GEM) models in melanoma research have enhanced our understanding of tumor biology and therapeutic response, but fail to mimic tumor progression in clinical settings as most lack spatial and temporal control of endogenous tumor initiation. To better mimic human disease, we have developed a novel tamoxifen application method that reproducibly induces local tumor formation on the mouse ear, and we have incorporated a tdTomato fluorescent reporter allele (tdTomatoLSL) into an existing tamoxifen-inducible GEM model of BRAFV600E/PTEN-null melanoma. The tdTomato allele serves as a visual marker of Cre recombination in endogenous melanocytes and allows disease progression to be followed through macroscopic and multiphoton intravital imaging. To investigate therapeutic effects at the cell level, we have performed imaging studies of primary tumors before and after treatment with a selective MEK1/2 inhibitor (MEKi), Trametinib, which is a FDA approved therapy for malignant melanoma. Our melanoma induction method enables longitudinal studies of drug response that ultimately give rise to fully resistant tumors (up to 90 days on MEKi). Our studies show that this treatment causes a striking morphological change of the tumor cells as early as 3 days post-treatment. After several weeks of continuous treatment, the remaining tumor cells are highly spatially correlated with bundled collagen structures detected by second harmonic signal, suggesting that cellular milieu strongly influences drug response. Interestingly, our current studies with MEKi suggest that association of tumor cells with collagen fibrils initially provides a protective effect against this drug, however once true resistance emerges, this dependency on extracellular matrix in the tumor stroma diminishes. Furthermore, in parallel with our imaging studies, we have performed molecular analysis of tumor response using Mib/MS and mRNA-seq. By coupling our imaging studies with transcriptome and kinome reprogramming analysis, we hope to identify properties that promote resistance. Using these novel approaches, we have developed a model that enables direct observation of endogenous tumor development, plasticity, and resistance to targeted therapy at the cell level in situ for the first time. Citation Format: Hailey E. Brighton, Steven Angus, Tao Bo, David Darr, Norman E. Sharpless, Gary L. Johnson, James E. Bear. Intravital imaging of endogenous BRAFV600E melanoma reveals plasticity of tumor response and resistance to trametinib over time. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4382.