Immune checkpoint signaling and cancer immunotherapy

Abstract

Immune checkpoint blockade therapy has become a major weapon in fighting cancer. Antibody drugs, such as anti-PD-1 and anti-PD-L1, demonstrate obvious advantages such as broad applicability across cancer types and durable clinical response when treatment is effective. However, the overall response rates are still unsatisfying, especially for cancers with low mutational burden. Moreover, adverse effects, such as autoimmune symptoms and tumor hyperprogression, present a significant downside in some clinical applications. These challenges reflect the urgent need to fully understand the basic biology of immune checkpoints. In this review, we discuss regulation of immune checkpoint signaling at multiple levels to provide an overview of our current understanding of checkpoint biology. Topics include the regulation of surface expression levels for known immune checkpoint proteins via surface delivery, internalization, recycling, and degradation. Upon reaching the surface, checkpoints engage in both conventional trans and also cis interactions with ligands to induce signaling and regulate immune responses. Novel therapeutic strategies targeting these pathways in addition to classical checkpoint blockade have recently emerged and been tested in preclinical models, providing new avenues for developing next-generation immunotherapies.

Fig. 1. Regulation of surface expression of PD-1, PD-L1 and CTLA-4.

a Fut8-mediated core fucosylation pathway is required for PD-1 surface expression. Internalized PD-1 is ubiquitinated by FBXO38 for proteasomal degradation and can also be recycled to surface with the help of TOX in liver cancer microenvironment. b STT3-catalyzed N-glycosylation stabilizes PD-L1 surface expression. P-S195-induced abnormally glycosylation of PD-L1 causes ERAD. Internalized PD-L1 is either sorted to the lysosome by HIP1R for degradation or recycled to the cell surface with the help of CMTM6/4. PD-L1 is ubiquitinated by different E3 ligases (HRD1, Cullin3-SPOP, β-TrCP and STUB1) under different contexts, and deubiquitinated by CNS5. Palmitoylation of PD-L1 by DHHC3 suppresses its mono-ubiquitination and lysosomal degradation. c Mgat1 mediates CTLA-4 N-glycosylation and surface retention. Trafficking of CTLA-4 to the cell surface relies on the TRIM/LAX/Rab8 complex and PLD/ARF1-dependent exocytosis. Rapid CTLA-4 internalization is mediated by AP-2 binding to the unphosphorylated YVKM motif. Internalized CTLA-4 is either degraded in the lysosome or recycled to cell surface by LRBA. CTLA-4 in TGN can also be delivered to the lysosome for degradation through AP-1 binding.

Fig. 2. Ligand binding and signal transduction of immune checkpoint receptors.

a PD-L1 and PD-L2 are ligands for PD-1. PD-1 recruits protein tyrosine phosphatase SHP2/SHP1 via phosphorylated ITSM/ITIM, which in turn inhibits both TCR and CD28 signaling. SAP inhibits SHP2 activity to suppress PD-1 signaling. Both PD-1 and CD80 interact with PD-L1 in cis to restrict its trans ligation of PD-1. b CTLA-4 competes with CD28 on binding with CD80/86 binding to inhibit CD28 signaling. The phosphorylated YVKM motif of CTLA-4 recruits SHP2 to inhibit RAS. CTLA-4 also inhibits AKT activity through PP2A. CTLA-4 in Tregs reduces CD80/86 on APCs by trans-endocytosis, which requires KxxKKR motif and PKCη. c TIM3 expresses in both T cells and innate immune cells. Four known ligands have been identified: Ceacam1, Galectin9, HMBG1, and PS. Galectin9 binds to glycosylated IgV domain of TIM3 in T cells. Ceacam1 exhibits both cis and trans interactions. Cis interaction of Ceacam1 with TIM3 is essential for TIM3 surface expression in T cells. In the absence of ligands, Bat3 binds to unphosphorylated Y256/263 in TIM3 cytoplasmic domain and recruits active Lck to deliver stimulatory signal in T cells. Interaction with Galectin9/Ceacam1 leads to phosphorylation of TIM3 Y256/263 and the subsequent abolishment of Bat3 binding, thus converting TIM3 from a stimulatory to an inhibitory molecule. TIM3 in DCs binds with PS and HMBG1 to regulate innate immunity. d LAG3 binds to MHC-II to inhibit CD4-dependent T cell function with its cytoplasmic domain. TME-derived Galectin3, LSECtin and FGL1 bind with LAG3 to inhibit T cell function, which requires the KIEELE motif in the LAG3 cytoplasmic domain. TCR signaling upregulates activity of ADAM10 and ADAM17, which cleave LAG3 at the extracellular domain to abolish its suppression of T cell signaling. e TIGIT and CD226 bind to the same ligands, CD112 and CD155. CD226 is a co-stimulatory receptor whereas TIGIT is a co-inhibitory receptor. TIGIT binds with CD112/CD155 with higher affinity than CD226 and inhibits the PI3K, MAPK and NF-κB pathways by recruiting SHIP1. f BTLA interacts with HVEM both in trans and cis. The cis interaction between BTLA and HVEM inhibits the trans-ligation of HVEM by LIGHT and thus inhibits HVEM stimulatory signaling triggered by LIGHT binding. ITIM and ITSM in BTLA recruit SHP1/SHP2 to inhibit both TCR and CD28 signaling.