Detection of functional tumor infiltrating T cells (TIL) positively correlates with improved prognosis and overall survival cancer patient. However, TIL are subjected to immune evasion and tolerance mechanisms that suppress defense against tumor growth. Therefore, TIL usually have a dysfunctional phenotype in solid tumor microenvironments. Such dysfunction includes decreased proliferation and progression through the cell cycle. Using the CT26 colorectal cancer model in mice, we readily detect T cells specific for the immunodominant tumor-associated antigen, AH1 (gp70423-431). To determine potential therapeutic targets, we compared gene expression profiles of tumor-specific T cells from the tumor and periphery. The results of this comparison highlight many known, hypothesized, and potential new mechanisms underlying TIL dysfunction. Gene set enrichment analysis of genes differentially regulated by TIL have significant overlap with those of exhausted LCMV-specific T cells published by Wherry in 2007. As expected, differentially expressed genes in TIL were also highly associated by pathway analysis with inhibition of T cell progression through cell cycle and decreased proliferation (Ingenuity). We have verified our microarray data by protein and functional analyses. TIL coexpress higher protein levels of multiple inhibitory receptors, produce less IFNγ, and progress less through the cell cycle in response to peptide stimulation. Studies by others suggest that E2F1 and E2F2 are involved in regulation of T cell progression through the cell cycle and proliferation. The following statistical analyses suggested members of the transcription factor family E2F are regulators of TIL dysfunction: 1) The top five transcription factor motifs found to be enriched in promoter regions of genes up-regulated by TIL are all variants for E2F1 (p < 1E-10, FDR < 1E-9, Fisher's exact test) and 2) activated E2F1 was predicted by upstream regulator analysis to have a key role in TIL gene regulation (Ingenuity). E2F has also been shown to be a key mediator of regulators of peripheral tolerance in the tumor environment such as PD1 and TGFβ (Patsoukis 2012, Lacerte 2008). E2F1 also regulates thymic negative selection, but is downstream of effector TCR stimulation through an undefined pathway (Jing Zhu 1999). Recently methyltransferase Prmt1 has been eloquently shown to be a posttranslational regulator of E2F1 negative regulation of cell cycle progression and apoptosis (Shunsheng Zheng 2013). Our data shows PRMT1 is induced in TIL. We hypothesize overexpression of E2F1/2 and PRMT1 contribute to the exhausted phenotype of TIL, particularly in T cells with high affinity for tumor antigen. We are currently testing our hypothesis with retrogenic mice that express TCRs of low or high affinity for tumor antigen, and combinatorial shRNA knockdowns of E2F1, E2F2, and PRMT1. Further defining TIL dysfunction at the molecular level may open a new frontier of effective therapeutic combinations.
This abstract is also presented as Poster A57.
Citation Format: Katherine A. Waugh, Sonia Leach, Jill E. Slansky. Determine a transcriptional mechanism novel to tumor infiltrating T cell dysfunction. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Immunology and Immunotherapy: A New Chapter; December 1-4, 2014; Orlando, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2015;3(10 Suppl):Abstract nr PR03.
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