A dual diacylglycerol kinase (DGK) alpha/zeta inhibitor augments the activity of human tumor infiltrating lymphocytes in in vivo and ex vivo models

Sep 28, 2026

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Endogenous or adoptively transferred tumor-infiltrating lymphocytes (TILs) often lose functional capacity as intrinsic inhibitory pathways are activated, restricting tumor control. This study blocks a key intracellular inhibitor, diacylglycerol kinase (DGK), using INCB165451, which targets DGKα and DGKζ. The inhibitor potentiates antitumor efficacy in a NSCLC mouse model, increases intratumoral infiltration, elevates inflammatory-response genes, and reduces TIL hypofunction. In human TILs from digests or precision-cut tumor slices, DGKi enhances proinflammatory cytokine and chemokine gene/protein expression after anti-CD3 stimulation and augments activation in EGFR/CD3 bispecific engager-stimulated tumor slices, supporting clinical translation.

LanguageEnglish

CategoryHealthcare

FormatPDF

Pages15

Reading Time38 min

AccessFree to download

Abstract

Introduction

Methods and Models

Results

In vivo adoptive T cell transfer

Ex vivo human TIL activation

Mechanistic readouts and cytokines

Keywords

Article Information

RESEARCH ARTICLE A dual diacylglycerol kinase (DGK) alpha/zeta inhibitor augments the activity of human tumor infiltrating lymphocytes in in vivo and ex vivo models Phurin Areesawangkita, b, Karen Pei-Yi Fonga, b, Emma Niemeyera, b, Yan Lia,c, Kelly Markowitza,c, Devora Delmana,c, Ryan Kraused, Justine Carle, Pat Feldmane, Lisa Trounge, Rodrigo Hesse, Xiaodi Rene, Cynthia Timmerse, Sunkyu Kime, Robert Brodyf, Sunil Singhald, Jarrod Predinad, Evgeniy Eruslanovd, Gregory Beattya,c, Bihui Melidosiane and Steven M. Albeldaa, b aCenter for Cellular Immunology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, USA; bDivision of Pulmonary and Critical Care Medicine, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, USA; cDivision of Hematology/Oncology, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, USA; dDivision of Thoracic Surgery, Department of Surgery, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA; eIncyte Corporation, Wilmington, Delaware, USA; fDepartment of Otorhinolaryngology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA ABSTRACT Endogenous or adoptively transferred tumor-infiltrating lymphocytes (TILs) often lose their functional capacity due to the activation of intrinsic inhibitory pathways, which then limits their ability to control tumor growth. In this study, we examined the effects of blocking a key intracellular inhibitory enzyme, diacylglycerol kinase (DGK) in human T cells, using a novel inhibitor (DGKi) called INCB165451 that blocks both DGKα and DGKζ, the two primary DGK isoenzymes that negatively regulate T cells through the diacylglycerol (DAG) signaling pathway. We first evaluated the effects of the DGKi in enhancing the efficacy of adoptive human T cell transfer in a non-small cell lung cancer (NSCLC) mouse model and found that the DGKi significantly potentiated anti-tumor efficacy through multiple mechanisms, including increased intratumoral T cell infiltration, upregulation of genes associated with inflammatory responses, and reduction of TIL hypofunction, as evidenced by enhanced cytokine production following ex vivo anti-CD3 antibody stimulation. We next studied the effects of the DGKi on human TILs derived from tumor digests or studied in situ in precision-cut tumor slices of both head and neck cancer and NSCLC patient samples. After stimulation of the TILs with anti-CD3 antibodies, we found that the DGKi enhanced gene and protein expression of proinflammatory cytokines and chemokines. Finally, we demonstrated that the DGKi could augment T cell activation in human tumor slices that were stimulated by an anti-EGFR/ anti-CD3 bispecific T cell engager (BiTE) . These data demonstrate strong activity of the DGKi in human TILs and highlight promising potential avenues for clinical translation. ARTICLE HISTORY Received 12 September 2025 Revised 20 November 2025 Accepted 18 December 2025 KEYWORDS T cells; tumor infiltrating lymphocytes; T cell hypofunction; diacylglycerol kinase; diacylglycerol kinase inhibitor; tumor digest; precision-cut tumor slices; adoptive T cell transfer; lung cancer; head and neck cancer Introduction Tumor-infiltrating lymphocytes (TILs) frequently lose their functional capacity, limiting their ability to control tumor growth. This dysfunction arises from various mechanisms 1,2 with a key contributor being the activation of intrinsic inhibitory pathways that normally prevent excessive immune responses and autoimmunity. These inhibitory regulators can be located on the T cell surface or within the cytoplasm. Monoclonal antibodies targeting surface immune checkpoint receptors, such as PD-1 and CTLA-4, have transformed cancer immunotherapy, offering substantial clinical benefit in many patient populations.3 Antibodies targeting additional surface receptors, like TIGIT and TIM3 are under

FAQs

Why do tumor-infiltrating lymphocytes (TILs) lose function?

TILs often become hypofunctional when intrinsic inhibitory pathways are activated, limiting their ability to control tumor growth.

What inhibitor was tested to block DGK in human T cells?

The study used INCB165451, a DGK inhibitor that blocks both DGKα and DGKζ, two primary DGK isoenzymes that negatively regulate T cells through DAG signaling.

How did the DGK inhibitor affect TIL activity in the study?

DGKi increased intratumoral T cell infiltration, enhanced inflammatory gene expression, improved cytokine production after anti-CD3 stimulation, and promoted proinflammatory cytokine and chemokine expression in human tumor slices.

Sep 28, 2026

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