Diacylglycerol kinase α inactivation is an integral component of the costimulatory pathway that amplifies TCR signals

Abstract

The arsenal of cancer therapies has evolved to target T lymphocytes and restore their capacity to destroy tumor cells. T cells rely on diacylglycerol (DAG) to carry out their functions. DAG availability and signaling are regulated by the enzymes diacylglycerol kinase (DGK) α and ζ, whose excess function drives T cells into hyporesponsive states. Targeting DGKα is a promising strategy for coping with cancer; its blockade could reinstate T-cell attack on tumors while limiting tumor growth, due to positive DGKα functions in several oncogenic pathways. Here, we made a side-by-side comparison of the effects of commercial pharmacological DGK inhibitors on T-cell responses with those promoted by DGKα and DGKζ genetic deletion or silencing. We show the specificity for DGKα of DGK inhibitors I and II and the structurally similar compound ritanserin. Inhibitor treatment promoted Ras/ERK (extracellular signal-regulated kinase) signaling and AP-1 (Activator protein-1) transcription, facilitated DGKα membrane localization, reduced the requirement for costimulation, and cooperated with enhanced activation following DGKζ silencing/deletion. DGKiII and ritanserin had similar effects on TCR proximal signaling, but ritanserin counteracted long-term T-cell activation, an effect that was potentiated in DGKα cells. In contrast with enhanced activation triggered by pharmacological inhibition, DGKα silencing/genetic deletion led to impaired Lck (lymphocyte-specific protein tyrosine kinase) activation and limited costimulation responses. Our results demonstrate that pharmacological inhibition of DGKα downstream of the TCR provides a gain-of-function effect that amplifies the DAG-dependent signaling cascade, an ability that could be exploited therapeutically to reinvigorate T cells to attack tumors.

DGK α and ζ Activities Control TH1 and TH17 Cell Differentiation

“…Previous studies have demonstrated that DGKα and ζ play crucial roles in T cell development, activation, anergy, and survival, and CD8 T cell-mediated anti-viral immune responses, iNKT cell development, regulatory T cell differentiation, and anti-tumor immune responses (27,(38)(39)(40)(41)(42)(43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53)(54). Additionally, DGKζ has been found to regulate B cell development (70), mast cell activation (71), TLR-mediated innate immunity (72), and NK cells (73).…”

“…DGKα and ζ, isoforms that express at high levels in T cells, have been demonstrated to inhibit the activation of both Ras-Erk and PKCθ-NFκB cascades as well as mTOR signaling (35)(36)(37). They regulate conventional αβT cell, iNKT cell, mucosal associated invariant T cell, and regulatory T cell development, negatively control T cell activation, regulate CD8 T cell mediated anti-viral responses and activation induced T cell death, promote T cell anergy, and inhibit anti-tumor responses (27,(38)(39)(40)(41)(42)(43)(44)(45)(46)(47)(48)(49)(50)(51)(52)(53)(54)(55). However, the role of DGKs in T H differentiation is unknown.…”

Subcellular Localization Relevance and Cancer-Associated Mechanisms of Diacylglycerol Kinases

“…Indeed, DGKα inhibition by Ritanserin induced cell death in glioblastoma stem cells and this was partially mediated by apoptosis [124]. Additionally, Ritanserin, as with other small molecule inhibitors of DGKα, also enhanced T cell signaling but failed to promote long-term T-cell activation [125].…”

Genome-wide CRISPR Screens in Primary Human T Cells Reveal Key Regulators of Immune Function

“…Many of the negative regulators are less well-annotated in the canonical TCR signaling pathway, although functions have been assigned to some. Diacylglycerol (DAG) kinases, DGKA (rank 17) and DGKZ (rank 1)negative regulators of DAG-mediated signalswere both found to restrain human T cell proliferation following stimulation (Arranz-Nicolás et al, 2018;Gharbi et al, 2011). The E3 ubiquitin-protein ligase, CBLB (rank 4) and its interacting partner, CD5 (rank 12), work together to inhibit TCR activation via ubiquitination leading to degradation of the TCR (Voisinne et al, 2016).…”