Genetic manipulation of OGT enhances NK cell-mediated cytotoxicity in tumor immunity

  • J Adv Res. 2025 Sep 4:S2090-1232(25)00678-2. doi: 10.1016/j.jare.2025.08.062.
Se-Chan Oh  1 ,  Bong Chan Jeon  2 ,  In-Hwan Jang  1 ,  Mi-Ran Song  3 ,  Heeyoun Hwang  4 ,  Dasom An  4 ,  Liu Yue  3 ,  Yuri Jung  3 ,  Yunhee Lee  5 ,  Seona Jo  6 ,  Sung-Kyun Park  7 ,  Tae-Don Kim  8
Affiliations
  • 1. Center for Gene and Cell Therapy, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea.
  • 2. Infectious Disease Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea; Department of Functional Genomics, KRIBB School of Bioscience, Korea University of Science and Technology (UST), Daejeon 34113, Republic of Korea.
  • 3. Infectious Disease Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea.
  • 4. Digital OMICs Research Center, Korea Basic Science Institute, Cheongju 28119, Republic of Korea.
  • 5. Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea.
  • 6. Center for Gene and Cell Therapy, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea; KRIBB School of Advanced Bioconvergence, University of Science and Technology (UST), Daejeon 34113, Republic of Korea.
  • 7. Infectious Disease Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea; Department of Functional Genomics, KRIBB School of Bioscience, Korea University of Science and Technology (UST), Daejeon 34113, Republic of Korea. Electronic address: [email protected].
  • 8. Center for Gene and Cell Therapy, Korea Research Institute of Bioscience and Biotechnology (KRIBB), Daejeon 34141, Republic of Korea; KRIBB School of Advanced Bioconvergence, University of Science and Technology (UST), Daejeon 34113, Republic of Korea; Department of Biopharmaceutical Convergence, School of Pharmacy, Sungkyunkwan University, Suwon 16419, Republic of Korea. Electronic address: [email protected].
PMID: 40914422 DOI: 10.1016/j.jare.2025.08.062
Abstract

Introduction: Natural killer (NK) cells are essential effectors in immune surveillance and Cancer Immunotherapy, but their function is often compromised by metabolic stress and environmental factors within the tumor microenvironment (TME). O-GlcNAcylation, a post-translational modification, regulates immune responses, yet its impact on NK cell function and therapeutic potential in immune cell-based therapies remains underexplored.

Objectives: This study investigates the effects of O-GlcNAcylation on NK cell-mediated cytotoxicity and its potential as a therapeutic target to enhance tumor immunity.

Methods: We investigated the impact of O-GlcNAcylation on NK cell cytotoxicity, focusing on its regulation under cytokine stimulation and pharmacological modulation. Mass spectrometry identified O-GlcNAc-modified proteins involved in NK cell cytotoxicity. NK92 cells were genetically engineered to delete the O-GlcNAc transferase (OGT) intronic splicing silencer (ISS) to ensure stable O-GlcNAcylation. The effects were evaluated under adverse TME conditions and in vivo tumor models. Gene expression analysis was performed to uncover the molecular networks underlying the observed effects.

Results: Cytokine stimulation and the O-GlcNAcase (OGA) inhibitor Thiamet G increased O-GlcNAc levels, enhancing NK cell cytotoxicity. Proteomic analysis identified key O-GlcNAc-modified proteins, including NK cell regulators and LRPPRC, which modulate NK function. Genetically engineered NK92 cells lacking the OGT-ISS region exhibited stable O-GlcNAcylation, preserving potent cytotoxicity under tumor-mimicking conditions and superior tumor-killing activity in vivo. Whole-transcriptome analysis of OGT-ISS-deleted NK cells revealed downregulation of TGF-β signaling and upregulation of Type I interferon signaling, as well as genes involved in cell adhesion and mobility, suggesting enhanced target recognition and cytotoxic function of NK cells.

Conclusion: Stabilization and enhancement of O-GlcNAcylation improve the target-killing capacity of NK cells while overcoming suppressive factors in the TME. These findings highlight advanced strategies, including genetic engineering of O-GlcNAc pathways, as potent approaches to augment NK-based immunotherapies against Cancer.

Keywords
Cancer immunotherapy; Cytotoxicity; NK cell; O-GlcNAcylation; OGT-ISS deletion.
Products