| Author: | Chan, Sze Man |
| Title: | Identification of IKBKB as a novel regulator of macrophage phagocytosis that suppresses antitumor immunity in hepatocellular carcinoma |
| Advisors: | Lee, Kin Wah Terence (ABCT) |
| Degree: | Ph.D. |
| Year: | 2026 |
| Subject: | Liver -- Cancer -- Immunotherapy Phagocytosis Macrophages Protein kinases Hong Kong Polytechnic University -- Dissertations |
| Department: | Department of Applied Biology and Chemical Technology |
| Pages: | xiv, 127 pages : color illustrations |
| Language: | English |
| Abstract: | Hepatocellular carcinoma (HCC) presents a significant clinical challenge owing to the high prevalence of primary and acquired resistance to current standard-of-care treatments, a problem often linked to an immunosuppressive TME. Cancer cells evolved to evade from clearance of macrophages by overexpressing the “don’t eat me” signal or antiphagocytic protein on the cell surface. Therapeutic strategies antagonizing the interaction of “don’t eat me” signals with macrophage-expressed receptors have been investigated in multiple cancer types, yet remain underexplored in HCC. Here, we aim to identify protein kinases governing resistance to phagocytosis in HCC. To this end, we employed an in vitro CRISPR-based kinome screen targeting 713 mouse kinase genes in a co-culture system in which mouse HCC cells (RIL-175) interacted with RAW 264.7 macrophages. Our analysis revealed that inhibitor of kappaB kinase beta (IKKβ), encoded by the IKBKB gene, emerged as a top sensitizer for macrophage-mediated phagocytosis, serving as a novel master regulator of anti-phagocytosis that facilitates immune evasion in HCC. Perturbation of Ikbkb significantly enhanced HCC cell phagocytosis and promoted macrophage activation under co-culture conditions. Depletion of Ikbkb suppressed HCC tumor growth, accompanied by increased in vivo macrophage phagocytosis and infiltration. The tumor-promoting role of Ikbkb through its anti-phagocytic function was further demonstrated by the observation that the growth-suppressive effect in Ikbkb knockout cells was offset by in vivo depletion of macrophages using clodronate liposomes. Mechanistically, Ikbkb facilitates evasion from macrophage-mediated phagocytosis by driving transcriptional activation of the anti-phagocytic checkpoints CD47 and PD-L1 via canonical NF-kB signaling. The immunomodulatory role of HCC cell-specific Ikbkb targeting was next assessed in immunocompetent murine models. Overexpression of Ikbkb in hepatocytes fosters tumor progression with a signature of compromised innate and adaptive immunities characterized by reduced infiltration and activation of innate immune cells, such as macrophages and dendritic cells, as well as dampened cytotoxic T cell activation and help T cells infiltration. Concordantly, cancer cell-specific Ikbkb depletion results in enhanced antitumor activities of both innate and adaptive arms, and remarkable tumor clearance. Interestingly, antigen cross-presentation was enhanced in splenic macrophages, suggesting systemic antigen spread and potentially accounting for the active immune surveillance of the adaptive arm and tumor clearance observed. Plausible therapeutic targeting was investigated using an IKKβ-targeting peptide, TAT-NEMOActpep. In cell models, treatment with TAT-NEMOActpep showed pro-phagocytic effects, resembling the results of gene depletion assays. Most critically, using the hydrodynamic tail vein injection (HTVI) model for hepatocarcinogenesis, which is known to generate HCC resistant to anti-PD-1 monotherapy, combining TAT-NEMOActpep with PD-1 blockade yielded effective tumor control without overt toxicity. Taken together, these findings establish IKKβ inhibition as an efficacious and bifunctional strategy to disrupt cancer cell defences and reprogram the immunosuppressive niche, providing a compelling rationale for co-targeting with immunotherapy to overcome treatment resistance in HCC. |
| Rights: | All rights reserved |
| Access: | open access |
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