Abstract B05: BRAF inhibition and cytokine therapy for melanoma: A novel rational combined approach
Authors: Romano Gabriele , Liang Roger , Liu Mingguang , Miller John , Costello James , Kwong Lawrence 
Published: May 2018

Abstract
Although the advent of BRAF inhibitors (BRAFi) plus MEK inhibitors (MEKi) has revolutionized melanoma therapy, most patients relapse within 16 months due to the persistence of minimal residual disease (MRD), a key obstacle in cancer therapy. Many efforts focus on combining this regimen with specific immune therapies, but clinical trials have so far uncovered no synergy and/or unacceptable toxicity. The critical knowledge gap that must be addressed in order to optimize such a combination is to identify the key drivers of the immune response to BRAFi as nodes for therapeutic synergy. To achieve this aim, we used our recently published model of melanoma (iBIP) to produce microarray data over a time course from early in BRAF extinction through to MRD and then tumor relapse. In the iBIP genetically engineered mouse model, doxycycline (dox) administration controls BRAFV600E expression. Interestingly, dox withdrawal resulting in BRAFV600E extinction is able to shrink the tumors, but not to make them completely disappear. As a result, in this model we have a minimal residual disease (MRD) of 5-20% of the original tumor volume. The microarray analysis confirmed that expected hallmarks of BRAFV600E extinction were present, resulting in BRAF and ERK pathway signature and a proliferation pathway signature decrease over time. Next, we detected a set of immune hallmarks with the following trend: acute and strong initial activation after BRAFV600E extinction, then a decrease through to MRD. These genes included markers of both CD4+ and CD8+ T cells, as well as NK cells and macrophages. As a result, we hypothesized that a strong immune infiltration, after an initial rapid induction in response to BRAFi, subsequently decreased and returned to baseline. This suggests that the MRD is immune suppressive or immune evasive.
To elucidate drivers of the immune signals, we first conducted a standard pathway analysis and identified cytokines as among the most-enriched immune genesets. We then performed a network analysis using a modification of our previously-described TRAP algorithm, and we identified CXCL9 (known to be involved in T cell mobilization) as the top candidate cytokine. Notably, CXCL9 is strongly induced within 8 hours of BRAFV600E extinction, but dissipates just prior to MRD, paralleling the more general immune signature. Even after iBIP tumors spontaneously relapse, CXCL9 continues to be low. Gene expression data obtained from human samples of BRAFi-responder patients confirm our mouse data showing CXCL9 higher expression in responding patients vs nonresponding. We have obtained promising preliminary results from a pilot experiment combining BRAF extinction with CXCL9 therapy, suggesting that CXCL9 can convert immunologically “cold” MRD back to “hot,” making them susceptible to immune rejection. As a result, in this work we present an approach that utilizes a confluence of in silico and in vivo data that guide the rational identification of a molecularly driven cancer treatment based on immune drivers with predicted molecular synergy.
Citation Format: Gabriele Romano, Roger Liang, Mingguang Liu, John Miller, James Costello, Lawrence Kwong. BRAF inhibition and cytokine therapy for melanoma: A novel rational combined approach [abstract]. In: Proceedings of the AACR Special Conference: Advances in Modeling Cancer in Mice: Technology, Biology, and Beyond; 2017 Sep 24-27; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(10 Suppl):Abstract nr B05.