Effectiveness of combination therapy based on ICIs with TME inhibitors or plant extracts in pre-clinical animal tumor models.
| Targets | Combination therapy | In vivo tumor model | Tumor growth and survival | Macrophage activity | Changes in the TME |
|---|---|---|---|---|---|
| Combination therapy of anti-PD-1/PD-L1 agents and TAM inhibitors | |||||
| MS4A4A [100] | Anti-MS4A4A mAb plus anti-PD-1 mAb | MC38-bearing C57BL/6 mice, CT26-bearing BALB/c mice (n = 5 per group) | Inhibited tumor growth (monotherapy); suppressed tumor growth and improved survival (combination) | An anti-MS4A4A monotherapy: decreased M2-TAM infiltration (F4/80+CD206+); downregulated M2-like markers (CD206) and immunosuppressive molecules (CD39, SIRPα); upregulated M1-like markers (iNOS, MHCII) | Reduced T cell exhaustion marker co-expression (PD-L1+LAG3+, PD-L1+TIM3+); enhanced CD8+ T cell effector function and proliferation (IFN-α, Ki67) |
| TREM2 [98] | CAR-T cells, secreted PD-1-TREM2 scFV | MC38-bearing C57BL/6J mice (n = 4) | Inhibited tumor progression; enhanced survival | Decreased percentages of M2-like TAMs (CD11b+Gr1+; F4/80+CD206+) | Increased CD8+ T cell infiltration; decreased MDSCs amount; enhanced perforin and granzyme B release; elevated intratumoral cytokines (IL-2, IL-15, TNF-α, IFN-γ) |
| SPHK1 [93] | SPHK1 inhibitors (PF543 or SKI II) plus anti-PD-1 Ab | MC38 and CT26 liver metastatic models in immunocompetent mice (n = 5–12 per group) | Partially repressed CRLM (monotherapy); enhanced metastasis inhibition; prolonged survival (combination) | Reduced total TAMs; decreased p-SPHK1+ TAMs | Increased intermediate exhausted CD8+ T cells with a moderate PD-1 level and PD-1lowCD8+ T cells expressing memory T cell markers (IL-7Ra and Ly6C) |
| CSF1R [102] | PLX3397 plus anti-PD-1 mAb plus anti-CTLA-4 mAb | C57BL/6J bearing subcutaneous MC38 mice | Compared with the monotherapy, PLX3397 showed a potent synergic effect when it was combined with an anti-PD-1 or an anti-CTLA-4 mAb | Decreased expression of macrophage markers CD206 and F4/80 (monotherapy PLX3397) | - |
| Combination therapy of anti-PD-1/PD-L1 agents and TME inhibitors | |||||
| A20 [95] | A20-knockdown plus, anti-PD-1 Ab | Metastatic BALB/c mouse model bearing A20-knockdown tumor cells (CT-26) (n = 4–8 per group) | Inhibited tumor growth in the lung, subcutaneous tumor growth; decreased number of tumor nodules in lung tissue; the overall survival was remarkably longer | Increased F4/80+ macrophages | Increased CD3+ and CD8+ T cells; increased amount of granzyme B+ immune cells |
| MIF [96] | anti-PD-1 Ab plus anti-MIF Ab | MC38-bearing (subcutaneously injected); C57BL/6J mice | Superior tumor rejection; reduction of tumor size | - | - |
| CD73 [97] | AB680 plus anti-PD-1 Ab | CT26-bearing BALB/c mice (n = 3 per group) | Tendency to reduce tumor growth compared to monotherapy | Increased the proportion of M1-like macrophages, no effect on M2-like macrophages (combination therapy); decreased the proportion of M2-like macrophages but had no effect on M1 macrophages (PD-1 blockade) | Trend towards increased CD8+/CD4+ ratio |
| Kinases [92] | Regorafenib plus anti-PD-1 Ab | Syngeneic murine MSS CT26 and MSI MC38 CC models | Sustained suppression of liver metastasis even after the end of treatment | Reduced F4/80+ and CD206+ macrophages; increased iNOS+ macrophages | Reduced number of Tregs under regorafenib treatment; increased intratumoral levels of IFN-γ, a pharmacodynamic marker of cytotoxic T cell activity under an anti-PD-1 therapy |
| WNT11/CAMKII [112] | KN93 plus anti-PD-1 Ab | C57BL/6 mouse liver metastases model with MC38 cells (n = 5 per group) | Decreased liver metastasis | Reduced CD206+ macrophage infiltration in liver metastasis | Increased infiltration of I-A/I-E+CD8+ T cells in liver metastasis; KN93 induced the expression of CXCL10 and CCL4 and reduced the expression of IL17D |
| Combination therapy of anti-PD-1/PD-L1 agents and epigenetic modulators | |||||
| HDAC [110] | Tucidinostat plus anti-PD-L1 Ab | CT26 subcutaneously injected into BALB/c or C57 BL/6 mice (n = 7 per group) | Tumor growth suppression; improved mice survival | Reduced the number of CD45+CD11b+F4/80+ TAMs; increased the proportion of CD45+CD11b+F4/80+/MHCII+ M1-like macrophages | Increased CD45+ lymphocytes count, proportion of tumor-infiltrating CD4+ T cells (CD45+CD3+CD4+) cells and CD8+ T cells (CD45+CD3+CD8+) cells; decreased expression of the exhaustion marker PD-1; increased serum level of CCL5 and IFN-γ |
| Class I HDACis (1, 2, 3, and 10), VEGFR [103] | Chidamide plus cabozantinib/regorafenib plus anti-PD-1 Ab | CT26-Bearing (subcutaneously injected) BALB/c mice | Inhibited tumor growth; increased survival rate; tumor shrinkage after discontinued treatment | Decreased amount of CD11b+ TAMs in tumors | Upregulated enrichment of IFN pathway gene signature and downregulated enrichment of the T cell gene signatures; reduced levels of polymorphonuclear-MDSCs in the tumor |
| Class I HDACis (1, 2, 3, and 10), VEGFR [103] | Chidamide plus cabozantinib/regorafenib plus anti-CTLA-4 Ab | CT26-Bearing (subcutaneously injected) BALB/c mice | Inhibited tumor growth; enhanced ORR | Suppressed macrophage/monocyte gene signatures | Enhanced T cell cytotoxicity |
| HDAC [111] | AVS100 plus anti-PD-1 Ab | CT26-bearing BALB/c mice (n = 6–7 per group) | An increase in responders from 35% in anti-PD-1 treatment alone to 80% in combination therapy; did not relapse after termination of the treatment; resistant to a subsequent tumor challenge | Increased IFN-TAMs; decreased regulatory and angiogenic TAMs | Up-regulation of genes involved in T cell activation, T cell effector function, and NF-κB signaling; increased expression of genes controlling IFN and NF-κB signaling (Stat4, Nfkb1, Rel, and Il12rb2), the inositol-3-phosphate/Akt pathway, and immune infiltration (Hcst, Itpr1, and Itgal1) with AVS100 treatment, independent of anti-PD-1 therapy |
| Combination therapy of anti-PD-1/PD-L1 agents and plant extract components | |||||
| [104] | Lobeline plus anti-PD-1 Ab | C57BL/6 mice bearing MC38 tumors (subcutaneously injected) (n = 6 per group) | Reduced tumor growth; upregulated SLURP1 expression in tumors; suppressed cancer cell proliferation | Increase in the number of intratumor CD45+CD11b+F4/80+ M1-like macrophages and decrease in the number of CD45+CD11b+CD206+ M2-like macrophages | Decreased Foxp3 protein levels; increased granzyme B levels |
| [113] | Astragaloside IV plus anti-PD-1 Ab | BALB/c mice bearing CT26 tumors (injected into the axillary fat pad) (n = 8) | Suppressed tumor growth more effectively than monotherapy | Astragaloside monotherapy: altered M2-like macrophage polarization toward M1-like (decreased CD11b+F4/80+CD206hi M2-like macrophages; increased CD11b+F4/80+MHCIIhi M1-like macrophages) | Increased the number of cytotoxic T cells; increased IFN-γ and IL12p70 secretion; decreased TGF-β secretion |
| [105] | SFIH plus anti-PD-1 Ab | MC38-bearing (subcutaneously injected), C57BL/6 mice (n = 7–8 per group) | Significant reduction in tumor volume compared with an anti-PD-1 Ab; no effect in the SFIH group | Increased CD11b+, F4/80+, MHCII+, CD206– M1-like macrophage infiltration; no changes in the CD11b+, F4/80+, MHCII–, CD206+ TAM population | Increased CD3+ and CD8+ T cells, release of granzyme B, expression of IFN-γ and CXCL9; decreased population of CD11b+, GR+ MDSCs, decreased expression of TGF-β1, TGF-β2, VEGFA, and IDO2; downregulated TGF-β signaling |
| [114] | CYP plus anti-PD-1 mAb | Xenograft models, subcutaneously injected: MC38-bearing C57BL/6 mice (n = 6) and CT26-bearing BALB/c mice (n = 3) | Combination therapy was more effective in reducing tumor volume and size (CYP alone had no significant anti-tumor effect); increased survival in mice | Decreased number of infiltrating macrophages, mainly CD206+ TAMs | Increased number of CD8+ T cells in the tumor; increased expression of granzyme B |
Ab: antibody; CAR-T: chimeric antigen receptor T; CCL: C-C motif chemokine ligand; CRLM: colorectal cancer liver metastasis; CTLA-4: cytotoxic T lymphocyte-associated antigen-4; CXCL: C-X-C motif chemokine ligand; CYP: Chinese yam polysaccharide; HDAC: histone deacetylase; ICIs: immune checkpoint inhibitors; IFN: interferon; IL: interleukin; mAb: monoclonal Ab; MDSCs: myeloid-derived suppressor cells; MHCII: major histocompatibility complex II; MIF: migration inhibitory factor; MSI: microsatellite instability; MSS: microsatellite stable; ORR: objective response rate; PD-1: programmed cell death-1; PD-L1: programmed cell death ligand-1; scFv: single-chain variable fragment; SFIH: sesquiterpene lactones derived from Inula helenium L.; SPHK1: sphingosine kinase; TAM: tumor-associated macrophage; TGF: transforming growth factor; TME: tumor microenvironment; TNF: tumor necrosis factor; Tregs: regulatory T cells; VEGF: vascular endothelial growth factor.