Ubiquitination in T-Cell Activation and Checkpoint Inhibition: New Avenues for Targeted Cancer Immunotherapy

Abstract

The advent of T-cell-based immunotherapy has remarkably transformed cancer patient treatment. Despite their success, the currently approved immunotherapeutic protocols still encounter limitations, cause toxicity, and give disparate patient outcomes. Thus, a deeper understanding of the molecular mechanisms of T-cell activation and inhibition is much needed to rationally expand targets and possibilities to improve immunotherapies. Protein ubiquitination downstream of immune signaling pathways is essential to fine-tune virtually all immune responses, in particular, the positive and negative regulation of T-cell activation. Numerous studies have demonstrated that deregulation of ubiquitin-dependent pathways can significantly alter T-cell activation and enhance antitumor responses. Consequently, researchers in academia and industry are actively developing technologies to selectively exploit ubiquitin-related enzymes for cancer therapeutics. In this review, we discuss the molecular and functional roles of ubiquitination in key T-cell activation and checkpoint inhibitory pathways to highlight the vast possibilities that targeting ubiquitination offers for advancing T-cell-based immunotherapies.

Keywords: E3 ligases; T cells; cancer immunotherapy; checkpoint inhibition; deubiquitinases; ubiquitination.

Figure 1

Mechanisms and functions of the ubiquitin system. (a) Enzymatic steps and enzymes involved in protein ubiquitination, a reversible and versatile post-translational modification. (b) Types of ubiquitin chains and their, thus far, identified cellular functions. Each ubiquitin chain has a different topology and is presumed to have distinct functions. The lysine 48 (K48)- and lysine 63 (K63)-linked polyubiquitin chains are abundant and well-studied; for the other types of chains, many known as atypical chains, cellular functions are starting to be revealed. Ub: ubiquitin; E1: ubiquitin-activating enzyme; E2: ubiquitin-conjugating enzyme; E3: ubiquitin ligase; DUB: deubiquitinase; AMP: adenosine monophosphate; ATP: adenosine triphosphate; PPi: inorganic pyrophosphate; K: Lysine; M: methionine; TGF-β: transforming growth factor-β. Illustration created using BioRender (biorender.com).

Figure 2

Ubiquitination in key T-cell activation pathways. T cells are activated upon antigen recognition by TCR complex (signal 1) and simultaneous engagement of co-stimulatory receptor such as CD28, 4-1BB, and ICOS (signal 2). These dual pathways integrate to trigger numerous signaling pathways that converge in the activation of NF-κB, NFAT, and AP-1 transcription factors that drive the genetic program for T-cell survival, proliferation, differentiation, and cytokine production. Various ubiquitin E3 ligases and deubiquitinases regulate these essential cascades using different ubiquitin chains, some of which activate (green chains), inhibit (purple chains), or degrade, via the lysosome or proteosome, (blue chains) the protein substrates. Ub: ubiquitin; DUB: deubiquitinase; AKT: serine/threonine-protein kinase/protein kinase B; AP-1: activator protein 1; BCL10: B cell lymphoma/leukemia 10; CARMA1: caspase recruitment domain-containing membrane-associated guanylate kinase protein-1; Cbl-b: casitas B-Lineage Lymphoma proto-oncogene B; CD40L: CD40 antigen ligand; CDC42: Cell division cycle 42; CDK: cyclin-dependent kinase; cIAP1/2: cellular inhibitor of apoptosis protein 1/2; CYLD: CYLD lysine 63 deubiquitinase; DAG: diacylglycerol; ERK: extracellular signal-regulated kinases; GITR: glucocorticoid-induced TNFR-related protein; GRAIL: gene related to anergy in lymphocytes protein; ICOS: inducible T-cell costimulator; IFNγ: interferon gamma; IKK complex: IκB kinase complex; IL2: interleukin 2; IP3: inositol trisphosphate; ITCH: itchy E3 ubiquitin protein ligase; I-κB: inhibitor of nuclear factor kappa B; KIP1: cyclin-dependent kinase inhibitor 1B; LAT: linker for activation of T cells; LCK: lymphocyte-specific protein tyrosine kinase; LUBAC: linear ubiquitin chain assembly complex; MALT1: mucosa-associated lymphatic tissue 1; MDM2: mouse double minute 2 homolog; NEDD4: neural precursor cell expressed developmentally down-regulated protein 4; NFAT: nuclear factor of activated T-cells; NF-κB: nuclear factor kappa-light-chain-enhancer of activated B cells; NRDP1: neuregulin receptor degradation protein-1 (also RNF41, ring finger protein 41); OTUB1: OTU domain-containing ubiquitin aldehyde-binding protein 1; OTUD7b: OTU deubiquitinase 7B; OTULIN: OTU deubiquitinase with linear linkage specificity; OX40: tumor necrosis factor receptor superfamily member 4 (also TNFRSF4); PIP2: phosphatidylinositol 4,5-bisphosphate; PKCθ: protein kinase C Theta; PLCγ1: phospholipase C-γ1; PTEN: phosphatase and tensin homolog; Rac1: rac family small GTPase 1; RhoGDI: Rho GDP-dissociation inhibitor; Roquin1/2: ring finger and CCCH-type zinc dinger somains 1/2; SCFSKP2: Skp1-Cullin-1-F-box (SCF) Cullin-Ring E3 ubiquitin ligase complex containing S-phase kinase associated protein 2 (SKP2); SLP76: lymphocyte cytosolic protein 2; TAB2: TGF-beta activated kinase 1 Binding Protein 2; TAK1: TGF-beta activated kinase 1 (also MAP3K7); TCR: T-cell receptor; TRAF1/2/5/6: TNF receptor associated factor 1/2/5/6; USP12/15/18/9X: ubiquitin specific peptidase 12/15/18/9 X-Linked, respectively; VAV1: vav guanine nucleotide exchange factor 1; ZAP70: ζ-chain associated protein 70. Illustration created using BioRender (biorender.com).

Figure 3

Ubiquitination in the CTLA-4 and PD-1/PD-L-1 checkpoint inhibitory receptors. Upon interaction with their cognate ligands on the cell surface of APCs or tumor cells, T-cell co-inhibitory receptors such as CTLA-4 and PD-1 impair T-cell activation to shut down the immune response and permit tissue repair. The ubiquitin system controls the function and stability of these receptors through degradative (blue) and non-degradative chains (purple), significantly impacting the outcome of these signaling events. Ub: ubiquitin; P+ (orange circle): phosphorylated residues; ARIH1: ariadne RBR E3 ubiquitin protein ligase 1; AKT: serine/threonine-protein kinase/ protein kinase B; Cbl-b: casitas B-lineage lymphoma proto-oncogene B; c-Cbl: casitas B-lineage lymphoma proto-oncogene C; CDK4: cyclin-dependent kinase 4; CDK5: cyclin-dependent kinase 5; CMTM4/6: CKLF-like MARVEL transmembrane domain containing 4/6; CSN5: COP9 signalosome complex subunit 5; CTLA-4: cytotoxic T-lymphocyte-associated protein 4; Cu: Copper ions; EGFR: epidermal growth factor receptor; FBXO38: F-box protein 38; GRAIL: gene related to anergy in lymphocytes; GSK3α: glycogen synthase kinases α; GSK3β: glycogen synthase kinases β; IL2: interleukin 2; ITCH: itchy E3 ubiquitin protein ligase; JAK: janus kinase; MARCH8: membrane-associated ring finger 8; OTUB1: OTU domain-containing ubiquitin aldehyde-binding protein 1; PD-1: programmed cell death protein 1; PD-L1: programmed death-ligand 1; PI3K: phosphoinositide 3-kinase; SPOP: speckle type BTB/POZ protein; STAT3: signal transducer and activator of transcription 3; STUB1: STIP1 homology and U-box containing protein 1; TRIM21: tripartite motif containing 21; USP7/22/9X: ubiquitin specific peptidase (USP) 7/22/9X-Linked, respectively; β-TrCP: beta-transducin repeat containing E3 ubiquitin protein ligase. Illustration created using BioRender (biorender.com).