Diacylglycerol kinases (DGKs) are a family of enzymes that metabolize the lipid diacylglycerol (DAG) to form phosphatidic acid. In T cells, two isoforms of DGK, DGKα and DGKζ, serve an important role in dampening signal transduction downstream of the T cell receptor (TCR), where they phosphorylate DAG generated by PLCγ1 after activation of proximal signaling molecules. DAG generated downstream of TCR binds to Protein Kinase C θ (PKCθ) and Ras Guanyl Releasing Protein 1 (RasGRP1), facilitating T cell activation. Elimination of either DGK isoform leads to increased generation of DAG and subsequent strengthened signaling downstream of RasGRP1 and PKCθ. Functionally, this results in formation of CD8+ T cells with enhanced cytokine production and proliferation after TCR stimulation. While mice or CD8+ T cells deficient in DGKα or DGKζ are both known to demonstrate enhanced immune responses to tumor, there has not been, to date, a direct comparison of the anti-tumor activity between mice lacking the two isoforms. We sought to determine which isoform of DGK more intensely attenuated anti-tumor immunity in order to identify which would be a better target for immune-based cancer therapies. We inoculated C57BL/6 mice with a variety of syngeneic tumor cell lines - MC38 colon carcinoma, B16F1 melanoma, and C1498 leukemia - and directly compared survival and tumor growth between mice deficient in DGKα or DGKζ in the presence or absence of anti-PD1 treatment. We found that DGKζ-/- mice more efficiently suppressed growth of subcutaneously implanted tumor cells when compared with DGKα-/- mice across the three model systems and further observed that the combination of DGKζ-deficiency and anti-PD1 were additive in tumor control. Additionally, relative to wildtype CD8+ T cells, we found that DGKζ-/- CD8+ T cells demonstrated qualitatively enhanced mRNA changes as assessed by RNA-seq analysis of cells after TCR stimulation. Pathway analyses from RNA-seq similarly indicated enhanced quantitative changes in DGKζ-/- T cells relative to DGKα-/- T cells, findings confirmed with in vitro biochemical and functional assays. Lastly, using CRISPR-based approaches to eliminate DGKα or DGKζ, we observed similar biochemical changes in human T cells when compared with T cells from gene-deficient mice. Our data, employing genetic approaches to modulate DGKα or DGKζ in both human and mouse models, suggests that DGKζ is a dominant regulator of TCR driven T cell activation and is consistent with the observed superior anti-tumor immune response.

Citation Format: Susan Wee, Junchen Gu, Cindy Wang, Carolyn Cao, Sandra Holzhauer, Heshani Desilva, Erin Wesley, Susan Tsai, Douglas Evans, Matthew Riese. Regulation of CD8+ T-cell function and antitumor activity by DGKα and DGKζ [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 936.