Update on Lymphocyte-Activation Gene 3 (LAG-3) in Cancers: From Biological Properties to Clinical Applications
Introduction
Immunotherapy targeting immune checkpoints has revolutionized cancer treatment, particularly through inhibitors of programmed cell death protein 1 (PD-1) and programmed cell death ligand 1 (PD-L1). However, the overall response rate to monotherapy remains limited, highlighting the need for more effective strategies. Lymphocyte activation gene 3 (LAG-3), an immune checkpoint receptor, has emerged as a promising target. Co-expressed with other inhibitory molecules, LAG-3 plays a critical role in immune suppression. Over the past five years, evidence has demonstrated the potential of LAG-3 blockade in enhancing anti-tumor immunity. This review provides a comprehensive update on the biological properties and clinical applications of LAG-3 in cancer.
Structure and Ligands of LAG-3
LAG-3 is a transmembrane protein composed of four immunoglobulin-like extracellular domains (D1–D4) and a cytoplasmic domain. Its extracellular region shares 20% amino acid identity with CD4, but its intracellular domain differs, leading to distinct functions. A connecting peptide between D4 and the transmembrane domain makes LAG-3 susceptible to cleavage by ADAM (a disintegrin and metalloproteinase domain-containing protein), producing soluble LAG-3 (sLAG-3). The cytoplasmic domain contains three motifs: a serine-based motif, a “KIEELE” motif, and a glutamic acid and proline dipeptide repeat (EP) motif, with the “KIEELE” motif primarily responsible for inhibitory activity.
The primary ligand of LAG-3 is major histocompatibility complex class II (MHC class II), with interactions mediated by a proline-enriched loop in D1. These interactions modulate immune cell proliferation, activation, apoptosis, and cytokine secretion. Other ligands include galectin-3, liver sinusoidal endothelial cell lectin (LSECtin), fibrinogen-like protein 1 (FGL-1), and α-synuclein preformed fibrils from neurons. Binding of LAG-3 to these ligands hampers anti-tumor T cell immunity, facilitating tumor immune evasion.
Regulation of LAG-3 at Epigenetic, Transcriptional, Post-Transcriptional, and Post-Translational Levels
LAG-3 expression is regulated at multiple levels, including epigenetic, transcriptional, post-transcriptional, and post-translational mechanisms. Epigenetic alterations, such as hypomethylation, have been reported in various cancers, including renal cell carcinoma (RCC), melanoma, breast cancer, and colorectal cancer. Hypomethylation of the LAG-3 promoter correlates with increased mRNA expression and immune cell infiltration, suggesting its potential as a predictive and prognostic biomarker.
At the transcriptional level, factors such as thymocyte selection-associated high mobility group box protein (TOX), nuclear factor of activated T-cells, and early growth response gene 2 enhance LAG-3 expression. Conversely, glycogen synthase kinase-3 reduces LAG-3 transcription by promoting Tbet expression. MicroRNAs in extracellular vesicles also modulate LAG-3 expression.
Post-transcriptional regulation involves N6-methyladenosine (m6A) modifications, with RNA demethylases like ALKBH5 and YTHDF1 influencing LAG-3 expression. Post-translationally, LAG-3 is degraded in lysosomes in the absence of antigen stimulation but translocates to the cell surface upon activation via protein kinase C signaling. Cleavage by ADAM10 and ADAM17 produces sLAG-3, which alleviates T cell inhibition and functions as a prognostic marker in multiple cancers.
Expression of LAG-3 and Its Role in Immune Suppression and Anti-Tumor Immunity
LAG-3 is expressed on various immune cells, including CD4+ and CD8+ T cells, natural killer (NK) cells, invariant NK T cells, plasmacytoid dendritic cells (pDCs), and B cells. Its overexpression in tumors is associated with immune regulation, treatment resistance, and patient survival.
On CD4+ T cells, LAG-3 is expressed following antigen stimulation and interacts with MHC class II to downregulate proliferation and cytokine secretion. The “KIEELE” motif is crucial for its inhibitory function, although its binding partner remains unidentified.
On CD8+ T cells, LAG-3 inhibits effector functions and inflammatory cytokine production. In RCC, CD8+ tumor-infiltrating lymphocytes (TILs) expressing LAG-3 are associated with disease progression. Inhibition of LAG-3, alone or with PD-1, enhances T cell activation and anti-tumor immunity.
LAG-3 is also expressed on CD4+CD25+ regulatory T cells (Tregs), suppressing dendritic cell maturation and immunostimulatory activities. It is specifically detected on IL-10-secreting T cells, which produce transforming growth factor-β3 to suppress B cell responses.
On other immune cells, LAG-3 expression does not affect NK cell activity but impairs IFN-γ production in NKT cells. On pDCs, LAG-3 regulates homeostasis and modulates T cell activity. B cells expressing LAG-3 suppress memory T cell formation through IL-10 production.
Associations of LAG-3 with Immunoregulatory Factors in Cancers
LAG-3 often co-expresses with other immune checkpoints, including PD-1, PD-L1, and cytotoxic T lymphocyte antigen 4 (CTLA-4). In various cancers, dual positive expression of LAG-3 and PD-1 is associated with an inflamed immunotype and IFN-γ release. Immune checkpoint inhibitors (ICIs) upregulate LAG-3 expression, suggesting a compensatory inhibitory mechanism.
LAG-3 is also associated with immune cells and inflammatory factors. Its expression correlates with activated CD8+ T cells, Tregs, myeloid-derived suppressor cells, and chemokines. In melanoma, LAG-3 co-expresses with CD163, a biomarker of M2-type tumor-associated macrophages, indicating a poor prognosis.
Correlation Between LAG-3 and Efficacy and Prognosis in Cancers
LAG-3 expression serves as a predictive biomarker for immunotherapy efficacy. In gastric cancer, high LAG-3 expression on TILs predicts longer progression-free survival (PFS). Conversely, elevated LAG-3 in NSCLC is associated with poorer PFS and resistance to PD-1 inhibitors. In breast cancer, increased LAG-3 correlates with ICI resistance.
Prognostically, higher LAG-3 expression in tumor tissues is linked to adverse outcomes in pancreatic cancer, soft-tissue sarcoma, and renal cell carcinoma. However, in ovarian cancer and blood cancer, high LAG-3 correlates with better survival. Contradictory findings in NSCLC, colorectal cancer, and breast cancer suggest that tumor origin, stage, and location influence the prognostic value of LAG-3.
Clinical Application of Targeting LAG-3 in Cancers
LAG-3-targeting agents, including monoclonal antibodies, sLAG-3, and bispecific antibodies, are being tested in clinical trials. Anti-LAG-3 monoclonal antibodies like relatlimab have shown efficacy in melanoma, with combination therapy (relatlimab plus nivolumab) significantly improving PFS.
sLAG-3, such as IMP321, activates antigen-presenting cells without inducing inhibitory signals. IMP321 vaccination induces durable anti-tumor immune responses, and combination therapy with pembrolizumab shows promise in NSCLC and melanoma.
Bispecific antibodies targeting LAG-3 and PD-1/PD-L1/CTLA-4, such as MK-4280A and MGD013, enhance immune stimulation. Preclinical studies demonstrate their potential to overcome tumor resistance, although their mechanisms require further investigation.
Conclusions
LAG-3 blockade enhances the efficacy of immunotherapy, particularly in combination with PD-1/PD-L1 inhibitors. Its expression on multiple immune cells and interactions with ligands play key roles in immune escape. However, the mechanisms of downstream inhibitory signaling and ligand-specific activation remain unclear. Further research is needed to optimize combination immunotherapy and reverse tumor resistance.
doi.org/10.1097/CM9.0000000000001981
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