Cancer Therapy: Preclinical The BRAF and MEK Inhibitors Dabrafenib and Trametinib: Effects on Immune Function and in Combination with Immunomodulatory Antibodies Targeting PD-1, PD-L1, and CTLA-4

Purpose: To assess the immunologic effects of dabrafenib and trametinib in vitro and to test whether trametinib potentiates or antagonizes the activity of immunomodulatory antibodies in vivo. Experimental Design: Immune effects of dabrafenib and trametinib were evaluated in human CD4þ and CD8þ T cells from healthy volunteers, a panel of human tumor cell lines, and in vivo using a CT26 mouse model. Results: Dabrafenib enhanced pERK expression levels and did not suppress human CD4þ or CD8þ T-cell function. Trametinib reduced pERK levels, and resulted in partial/transient inhibition of T-cell proliferation/expression of a cytokine and immunomodulatory gene subset, which is context dependent. Trametinib effects were partially offset by adding dabrafenib. Dabrafenib and trametinib in BRAF V600E/K, and trametinib in BRAF wild-type tumor cells induced apoptosis markers, upregulated HLA molecule expression, and downregulated certain immunosuppressive factors such as PD-L1, IL1, IL8,NT5E, andVEGFA. PDL1 expression in tumor cells was upregulated after acquiring resistance to BRAF inhibition in vitro. Combinations of trametinib with immunomodulators targeting PD-1, PD-L1, or CTLA-4 in a CT26 model were more efficacious than any single agent. The combination of trametinib with anti–PD-1 increased tumor-infiltrating CD8þ T cells in CT26 tumors. Concurrent or phased sequential treatment, defined as trametinib lead-in followed by trametinib plus anti–PD-1 antibody, demonstrated superior efficacy compared with anti–PD-1 antibody followed by anti–PD-1 plus trametinib. Conclusion: These findings support the potential for synergy between targeted therapies dabrafenib and trametinib and immunomodulatory antibodies. Clinical exploration of such combination regimens is under way. Clin Cancer Res; 21(7); 1639–51. 2015 AACR. Introduction Immunotherapies and targeted therapies have distinctly different mechanisms of action and have both been shown to be efficacious in patients with advanced cancers (1, 2). It is expected that combinations of both modalities may create synergies with increased benefit for patients with cancer. To enable such combinations, it is critical to determine how targeted therapies affect immune function in the tumormicroenvironment andperipheral systems. Immunogenic cell death, characterized by secretion of cell damage–associated hallmark molecules consisting of calreticulin (CRT), HSP70 and 90 proteins, HMGB1, and ATP, increased expression of tumor antigens and HLA molecules, and decreased expression of immunosuppression factors are desirable features for potential immune sensitization (3, 4). These effects may allow targeted agents to not only directly inhibit tumor growth, but also further enhance immune response by immunotherapy, through either tumor cell intrinsic or extrinsic immunomodulatory mechanisms, thus making the cancer therapy more effective and durable. Inhibition of oncogenic MAPK signaling by dabrafenib, trametinib, or the combination of dabrafenib and trametinib has been an effective strategy and approved for the treatment of metastatic melanoma tumors bearing BRAF V600E and V600K mutations (5). The first generation of checkpoint immunomodulatory antibodies targeting either CTLA-4 or PD-L1/PD-1 has demonstrated impressive clinical activity resulting in durable responses in subsets of patients with various cancer types (1, 6, 7). BRAF inhibitors, such as vemurafenib, have been shown to increase immune response and efficacy in combination with immunomodulators in preclinical models (8, 9). However, MEK inhibitors, including trametinib, have been reported to be immunosuppressive in vitro, which has limited the in vivo assessment of MEK inhibitor combinations with immunotherapies (10). In this study, we assessed the immunologic effects of dabrafenib and trametinib at clinically relevant exposures on both immune and Immuno-Oncology and Combination DPU, GlaxoSmithKline, Collegeville, Pennsylvania. Molecular Medicine Unit, Oncology R&D, GlaxoSmithKline, Collegeville, Pennsylvania. Statistical Science, GlaxoSmithKline, Collegeville, Pennsylvania. Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). L. Liu and P.A. Mayes contributed equally to this article. Corresponding Author: Axel Hoos, Immuno-Oncology and Combination DPU, GlaxoSmithKline, 1250 S. Collegeville Road, Collegeville, PA 19426. Phone: 610-427-3733; E-mail: [email protected] doi: 10.1158/1078-0432.CCR-14-2339 2015 American Association for Cancer Research. Clinical Cancer Research www.aacrjournals.org 1639 on April 14, 2017. © 2015 American Association for Cancer Research. clincancerres.aacrjournals.org Downloaded from Published OnlineFirst January 14, 2015; DOI: 10.1158/1078-0432.CCR-14-2339