4339527

Discovery of a covalent inhibitor of diacylglycerol kinase (DGKα) for immunotherapy applications

Date
August 20, 2025

Diacylglycerol kinases (DGKs) regulate T cell receptor (TCR) signaling by converting diacylglycerol (DAG) to phosphatidic acid (PA), thereby attenuating downstream DAG signaling. Among DGK isoforms, DGKα and DGKζ are predominantly expressed in T cells, where they serve as metabolic checkpoints of DAG signaling. Several DGK inhibitors have been reported but the DGK isoform and proteome-wide selectivity of these agents remains unknown. To address this challenge, we report the deployment of a chemoproteomic platform to identify ligandable sites and design isoform-selective DGK inhibitors, with a focus on DGKα. We synthesized a series of covalent inhibitors and assessed their biochemical activity in vitro. Our findings demonstrate that these inhibitors exhibit potent and selective inhibition of DGKα over DGKζ. Cellular assays showed that our lead DGKα inhibitor enhanced Jurkat T cell activation in response to anti-CD3 and anti-CD28 stimulation. Additionally, we provide preliminary evidence for stimulatory activity in CD8+ T cells. Using the PILS-Nir1 PA biosensor and ectopic DGKα localized to mitochondria, we confirmed dose-dependent inhibition of PA production, consistent with our lipidomics results showing increased DAG and reduced PA levels in Jurkat T cells. This study underscores the potential of chemical biology approaches for the discovery of isoform-selective small molecule inhibitors targeting DGKα, offering a foundation for future therapeutic strategies to modulate T cell function through selective DGK inhibition.

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