Abstract 958: Treatment with a selective inhibitor of BRAFV600E increases melanocyte antigen expression and CD8 T cell infiltrate in tumors of patients with metastatic melanoma
Authors: Wargo Jennifer A. , Cogdill Alex , Dang Ping , Gupta Ridhi , Piris Adriano , Boni Andrea , Garber Haven R. , Ott Harald , Newton Lindsay P. , Flaherty Keith T. , Lawrence Donald P. , Tsao Hensin , Fisher David E. 
Published: June 2012

Abstract
Targeted therapy against oncogenic BRAF (BRAFV600E) is a promising new therapeutic approach for the treatment of melanoma with up to 80% of patients with stage IV melanoma responding to treatment. Despite these results, the median duration of response is only 8.5 months. Investigators propose combining this therapy with other targeted agents to address redundancy and signaling through different oncogenic pathways in hopes to improve the durability of response. An alternate approach is to combine BRAF-targeted agents with immunotherapy based on evidence that oncogenic BRAF contributes to immune escape. We recently reported data supporting this hypothesis, showing that treatment of melanoma cell lines with MEK or selective BRAFV600E inhibition results in increased expression of melanocyte differentiation antigens (MDAs) which is associated with enhanced recognition by antigen-specific T cells.
The purpose of these studies was to validate these findings by assaying expression of MDAs in patients with metastatic melanoma treated with BRAFV600E inhibition. We also sought to test immune cell infiltrate in tumors to analyze host immune response.
Patients with metastatic melanoma undergoing treatment with a selective BRAFV600E inhibitor were consented on a tissue procurement protocol approved through the IRB at the Massachusetts General Hospital. Patients underwent treatment with the BRAFV600E inhibitor (PLX4032) per protocol (960 mg given twice daily). Tumor biopsies and blood draws were performed pre-treatment and 10-14 days after initiation of treatment. RNA was harvested and expression of MDAs was assayed by quantitative RT-PCR. The level of MITF, the master transcriptional regulator of melanocyte which controls MDA expression, was also assayed. Immunohistochemistry was performed on formalin-fixed, paraffin embedded tissue for MDAs, as well as CD4 and CD8. The effect of BRAFV600E inhibition on T lymphocyte function was analyzed by assaying fold-expansion and viability in blood samples collected before and during treatment.
Treatment with a selective inhibitor of BRAFV600E resulted in significantly increased levels of MDAs (up to 95-fold) which was associated with higher levels of MITF. In addition, CD8 T cell infiltrate also increased after treatment. Of note, T cell function was preserved after treatment with a specific inhibitor of BRAFV600E.
These data provide in vivo correlates to our in vitro findings that selective BRAFV600E inhibition enhances MDA expression in melanoma and suggest a possible improved host immune response. The studies provide further support for potential synergy between BRAF-targeted therapy and immunotherapy in the treatment of 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 958. doi:10.1158/1538-7445.AM2011-958