Antigen presentation in the body is often due to an insufficient number of DCs, which results in a failure to cause more CD8⁺ T cells to infiltrate the tumor (Gardner et al., 2020). Low immunogenicity and weak immune response limit abscopal effects in the treatment of metastatic tumors. Immunogenic cell death (ICD) promote the recruitment of DCs in metastases. Min et al. (2017) developed several antigen-capturing nanoparticles (ACNPs) based on poly (lactic-co-glycolic acid) (PLGA). Research suggests that positively charged particles were more likely to be captured by DCs. ACNPs delivered TAAs to APCs and promoted DCs trafficking to TDLNs to enhanced antitumor immune responses. In addition to TAAs, high-mobility group box 1 (HMGB1), damage-associated molecular patterns (DAMPs), and histone proteins have also been shown to promote the recruitment of DCs (Ren et al., 2018; McCaw et al., 2019; Wang and Zhang, 2020). Natural compounds often act as carriers to transport these substances around tumors. Because of its large surface area, large pore size, multifunctional surface modifiability, and biodegradability, silica has been widely used for drug delivery (Nguyen et al., 2020a). Deng et al. (2020b) developed silica mesoporous nanotechnology for the delivery of nanovaccines for metastatic cancer immunotherapy. The silica nanoparticles enhance the recruitment of DCs. However, whether the nanoparticles trigger the abscopal effect has not been verified.

Tumor cells are surrounded by a hypoxic environment, which is conducive to tumor cells escaping attack by the immune system (Fu and Xiang, 2020). Therefore, increasing the amount of oxygen in the tumor environment is an effective way to induce ICD and activate DCs to produce powerful anti-tumor effects. Gold nanocages (AuNCs) have attracted considerable attention because of their hollow porous structures and controllable optical performance. Liang et al. (2018a) encapsulated manganese dioxide with AuNCs (AuNC@MnO ₂) for targeting tumor-associated macrophages (TAMs) for oxygen production. AuNC@MnO ₂ was prepared by mixing AuNC and polyethylene glycol (PEG) in a potassium permanganate solution by magnetic stirring. With an average size of 91 nm and a surface charge of + 5.6 mV, AuNC@MnO₂ had the advantage of reactive oxygen species (ROS) responsiveness to generate oxygen around the tumor. The nanocages elicited ICD by provoking dying tumor cells to induce enhanced exposure and release of DAMPs, such as calreticulin, adenosine triphosphate (ATP), and HMGB1. DAMPs then promoted the recruitment of DCs and increased the antigen engulfment and presentation, followed by prominent activation of effector cells (e.g., CD8 T cells, CD4 T cells, and NK cells) for inhibition of primary tumor and lung metastases. AuNC@MnO ₂ combined with photodynamics up-regulated the expression of CD80 and CD86 up to 7.5% and reduced the number of lung metastases in tumor-bearing mice (Liang et al., 2018a). Yang et al. (2020) explored PEG cationic peptides to self-assemble into pH-responsive nanovesicles. PEG nanovesicles with a hydrodynamic diameter of 55 nm. In vitro experiments showed that PEG nanovesicles were endocytosed by tumor cells within 4 h. Encapsulation of photosensitizers into PEG nanovesicles generated ROS within tumors to induce tumor cell death. In tumor-bearing mice, PEG nanovesicles significantly inhibited the growth of primary and distant tumors within 17 days. Human serum albumin (HAS) shows natural biocompatibility, excellent stability, and positive tumor targeting ability. Chen et al. (2018) utilized reduced HAS and hemoglobin (Hb) through intermolecular disulfide hybridization to obtain hybrid protein oxygen carrier (HPOC) nanoparticles. The advantage of HPOC was that Hb’s own oxygen-carrying ability provided oxygen to the hypoxic environment of tumor cells and improved the treatment efficiency. In another study, Zhou et al. utilized maleimide liposomes to deplete intracellular glutathione. Liposomes remodeled TAMs to promote DCs recruitment (Zhou et al., 2020b).

CCL21 is a chemokine that promotes the recruitment of DCs (Hu et al., 2020). Owen et al. developed a hydrogel made of PEG-polylactic acid (PLA) for carrying CCL21 to promote DCs recruitment (Fenton et al., 2019). The advantage of hydrogel was that after injection into the mouse, CCL21 was released within 48 h to recruit DCs to the injection site. In another study, protamine sulfate loaded with seaweed shown high loading rate of CCL (Poelaert et al., 2020). Protamine sulfate acts as a polycation, stabilizing alginate nanoparticles into polyelectron complexes. Intratumoral injection of nanoparticles caused neuroblastoma to decline and improved the long-term immune effect of mice (Poelaert et al., 2020). However, no instances of the abscopal effect being triggered by CCL21 loaded on nanomaterials have yet been reported. We believe that nanomaterials to induce CCL21 to enter the metastatic tumor site to recruit DC is a promising method for metastatic tumor treatment.

Immature DCs cause T cells to be unresponsive to tumors and lead to immune tolerance (Dudek et al., 2013). Therefore, the maturation of DCs is crucial to achieving the abscopal effect in immunotherapy. Because of the presence of transforming growth factor-β (TGFβ) and interleukin 10 (IL10), the number of DCs in tumor patients is abnormal (Brown et al., 2001). Toll-like receptors (TLRs) are an important class of protein molecules involved in non-specific immunity. TLR9 is mainly expressed in B cells and plasmacytoid dendritic cells (pDCs) (Krieg, 2007). The activation of TLR9 on pDCs has a variety of effects, such as the expression of Interferon-γ (IFN-γ), Th1-type cytokines (tumor necrosis factors-α, IL-2), TNF-related apoptosis-inducing ligand (TRAIL), and co-stimulatory molecules (CD80, CD86) (Suek et al., 2019). TLR7 and TLR9 have been proven to be related to the maturity of DCs (Leong et al., 2019).

Natural polysaccharides are polymers with good biological safety and various biological functions (Zhu et al., 2016; Yu et al., 2018). Natural polysaccharide nanoparticles carrying TLR agonists caused more DCs activation (Pang et al., 2019). Although the application of TLR agonists has a good effect, it once caused obvious inflammation. Darling et al. (2020) explored acetic anhydride nanoparticles with a mixture of 3,6-dioxaoctante (CPTEG) and 1,6-bis (p-carboxyphenoxy) hexane (CPH) by a molar ratio of 20:80. The average particle size of the nanoparticles was approximately 200 nm. Acetic anhydride nanoparticles provided antigen delivery and DCs activation while avoiding the extensive inflammatory response usually associated with traditional adjuvants (Darling et al., 2020). Nanomaterials carrying TLR agonists have been reported to enhance DCs activation and trigger the abscopal effect (Cano-Mejia et al., 2020; Ni et al., 2020; Perry et al., 2020; Luo et al., 2021). For example, as a class of molecular nanomaterials, cationic nanoscale metal–organic frameworks (nMOFs) are suited for biomedical applications because of their crystallinity, tuneability, and porosity (Ni et al., 2020). nMOFs had emerged as novel nanophotosensitizers for photodynamic therapy (PDT) with high photosensitizer (PS) loadings, enhancing the abscopal effect of PDT triggering. Prussian blue (PB) also shows good biocompatibility and is widely used in biomedicine. Rohan et al. developed PB nanoparticles for the delivery of CpG and promoted the maturation of DCs in neuroblastoma (Cano-Mejia et al., 2020). PB nanoparticles alter the presence of immunomodulatory receptors and ligands on the treated tumor cells and generates an abscopal effect. PB exhibits pH-dependent biodegradability, mitigating concerns over long-term persistence and toxicity within the body (Cano-Mejia et al., 2020). The study found that the local release of ROS from the tumor also promoted the maturation of DCs. Wang et al. (2021a) devoloped light-activable immunological adjuvant (LIA) loaded with chlorin e6, after being internalized by tumor cells in a mouse model of breast cancer. LIA produced ROS under light to induce tumor cells to released antigens and also promoted DCs maturation.

Radiotherapy (RT) can break the DNA double bond of tumor cells, release TAAs, and cause tumor cells to produce ICD. RT has been widely used clinically. However, RT is normally used for localized treatment rather than to treat metastatic cancer, and it is not safe for certain areas of the body. Chen et al. explored cisplatin loaded within PLGA-graft-methoxy PEG complex nanoparticles (CDDP-NPs) (Figure 1) (Wang et al., 2021b). Platinum crosslinks the DNA double strand, reducing the stability of DNA by a process called DNA platination. This nanomaterial enhanced the body’s tolerance to radiation doses and caused more DNA damage in tumor cells. CDDP-NPs resulted in greater DNA platinization compared to CDDP at 24 h (51.02 × 10^–3 pg Pt per ng DNA), 48 h (38.80 × 10^–3 pg Pt per ng DNA), and 72 h (14.24 × 10^–3 pg Pt per ng DNA) (Figure 1B). Figure 1B The nanoparticles extended the residence time of platinum-based chemotherapeutics around the tumor but did not damage the cells of the whole body (Figure 1C). The combined application of cisplatin nanoparticles and radiotherapy increased the expression of CD80 in distant tumors (Figure 1D), boosted the abscopal effect and delayed the progression of primary and distant tumors (Figures 1E,F). However, CDDP-NPs induced hematopoietic suppression and peripheral cytopenias (Wang et al., 2021b). This may be detrimental to the application of CDDP. Therefore, further research is needed to reduce its side effects.

CD40/CD40L is another pathway that causes the maturation of DCs. CD40 increase the expression of immunoglobulin superfamily co-stimulatory molecules such as CD80 and CD86 (Vonderheide, 2020). CD40 signaling occurs through two adaptor proteins, TNFR-related factor (TRAF) and Jak family kinase 3 (JAK3). These proteins activate various signaling pathways, including MAPK, PI3K, and NF-kβ, which in turn cause DCs to mature. It has been confirmed that CD40 agonists boost abscopal effects by activating DCs (Wood et al., 2020). Although CD40 affects the maturation of DCs, CD40 stimulation alone does not enable mice to obtain long-term immunity but rather produces ineffective cytokines and specific T cells (Lau et al., 2020). The combination of CD40 antibodies and TLR agonists is generally more effective. Liu et al. developed nanofluidic drug-eluting seeds (NDES) for the delivery of CD40 antibodies and TLR agonists into tumors (Liu et al., 2020). The nanochannel of NDESs was 150 nm, and intratumoral injection of NDESs continuously and slowly released CD40 as long as 14 days. In combination with external radiation therapy, NDESs enhanced the abscopal effect and reduced the chance of metastasis by promoting the maturation of DCs.

In conclusion, some nanomaterials targeting DCs exert anti-tumor effects. Nanomaterials located with TLR agonists not only treat existing tumors but also play a role in preventing tumor metastasis. Not only that, nanomaterials have been proved to good biological tissue compatibility and targeting. Many effects, such as regulate DC maturation and activation, have been shown to exist simultaneously. These findings have opened up new avenues for progress in tumor treatment.

Immune checkpoints are inhibitory regulatory molecules in the immune system that are essential for maintaining self-tolerance and preventing autoimmune reactions. Immune checkpoints expressed on immune cells inhibit the function of immune cells so that the body does not produce enough effective anti-tumor immune responses. The immune checkpoint blockade (ICB) enhances anti-tumor activity by disrupting inhibitory T cells signaling (Shi et al., 2020). Programmed cell death protein 1 (PD-1) is an important immunosuppressive transmembrane protein expressed on the surface of T cells. PD-L1 is a ligand of PD-1. The combination of PD-1 on the surface of T cells with PD-L1 will transmit inhibitory signals and reduce the proliferation of CD8⁺ T cells in lymph nodes. One of the important mechanisms by which tumor cells evade the attack of immune cells is by expressing PD-L1 (Han et al., 2020). Metastatic cancer cells express more PD-L1 than normal tissue cells (Eckert et al., 2018). Blocking the PD-1/PD-L1 signaling pathway can thus enhance the killing of metastatic tumors by CD8⁺ T cells. In addition, cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) is a leukocyte differentiation antigen, a transmembrane receptor on T cells, and it shares the B7 molecular ligand with CD28. CD28 is a transmembrane protein on the surface of T cells. The APC surface expresses B7.1 (CD80) and B7.2 (CD86) ligands. The combination of CD28 and B7 molecular ligands can activate T cells. CTLA4 has a structure homologous to CD20 that can compete with CD28 for B7 ligands, resulting in insufficient T cell activation (Hosseini et al., 2020). Although ICB has achieved good results, it only works on tumor cells that have been pre-infiltrated by T cells (Spranger and Gajewski, 2018). In metastatic tumors, insufficient T cell infiltration has become a major obstacle to treatment. Therefore, how to induce the infiltration of T cells into metastatic TME and increase T cells’ activity have emerged as key problems in the pursuit of curing metastatic cancer.

Blocking immune checkpoints enables the immune system to recognize and attack metastatic tumor cells. However, the application of ICB alone cannot trigger the abscopal effect to eliminate metastases. The body needs to activate the immune system to induce tumor cells to produce ICD. Nanoparticles induce ICD to expose the tumor to more TAAs when nanoparticles enter the periphery of the tumor. With the application of ICB, more T cells can infiltrate the tumor. Huang explored a celastrol (CEL) nanoemulsion (Qiu et al., 2021). CEL is recognized for its anti-inflammatory effects via suppressing macrophage M1 polarization. In addition, nanoparticles caused an inflammatory response, promoted the recruitment of NK cells in metastatic tumors, and activated tumor-specific T cells to control metastatic tumors. In an experiment involving intravenous injection of CEL nanoemulsion and intraperitoneal injection of anti-PD-L1 in tumor-bearing mice, the CEL nanoemulsion/anti-PD-L1 treatment exhibited a good killing effect, inducing nearly 90% of the total cell apoptosis in treated tumors, with nearly 15-fold or 10-fold more apoptotic cells compared to the control, and anti-PD-L1-treated tumors. After the application of anti-PD-L1, the growth of the primary tumor and the distant tumor was significantly inhibited. Instead of delivering anti-PD1 directly, experimental mice are usually injected intraperitoneally with anti-PD1 to block ICB. Nanoparticles can synergize with ICB to induce stronger abscopal effects after intravenous injection. For examples, upconversion nanoparticles (UCNPs), ROS-sensitive lipid-polymer hybrid nanoparticles, PB nanoparticles and nanoliposomes loaded with pheophorbide A (PhA) have been shown to produce long-term immune memory effects and further enhance the ICB therapeutic outcome (Xu et al., 2017; Hu et al., 2019; Zhou et al., 2020c; Hwang et al., 2020). Tandem peptide nanocomplex (TPNC) and copper chalcogenide nanoparticles carrying CpG can also enhance the therapeutic effect of ICB (Buss and Bhatia, 2020; Cao et al., 2020).

Indoleamine 2, 3-dioxygenase (IDO) is one of the body’s main oxygenases for tryptophan metabolism. It can reduce cellular energy levels, prevent immune cell maturation, and induce T cell apoptosis (Heidari et al., 2020). IDO is the rate-limiting enzyme that metabolizes tryptophan to kynurenine in the body, which regulates the level of metabolism in the body and leads to immune tolerance in the body. IDO is expressed in the cytoplasm of a variety of cells in the TME, DLNs, and peripheral blood. IDO mainly affects the anti-tumor effect of T cells in the following ways. 1) The expression of IDO in tumors is related to the expression of forkhead box P3 (FoxP3) positive T-regs and monocytes in tumor infiltration, and these cells will negatively regulate CD8⁺ T cells in primary tumors and metastases (Meireson et al., 2020). 2) IDO down-regulates the expression of T cells receptors and inhibits T cells proliferation. 3) IDO also affects the function of T cells by reducing the expression of CD107a and granzyme B. 4) IDO also promotes the production of tumor blood vessels (Prendergast et al., 2018). 5) IDO is closely related to the expression of PD-1 and PD-L1 (Dill et al., 2018; Ye et al., 2018). Recent studies have confirmed that the combination of CTLA-4 and CD80/CD86 on APC can induce IDO expression (Walker and Sansom, 2015). IFN-γ produced by T cells and NK cells induces TRC to enter a dormant state through the IDO-Kyn-AhR-p27 cascade, thereby allowing tumor cells to escape immunity (Liu et al., 2017). Therefore, IDO inhibitors have become a hotspot in tumor immunotherapy.

Most IDO inhibitors cannot function effectively due to their short half-life and poor tumor accumulation (Kuang et al., 2018). Chen et al. (2020) encapsulated indocyanine green (ICG), a photosensitizer, into chitosan nanoparticles (ICG-NPs) (Figure 2A). The zeta potential of ICG-NPs was 47.2 ± 0.3 mV, and the particle size was approximately140 nm. ICG-NPs delivered photosensitizers into tumor cells and effectively regress 80% of tumors in melanoma mice. Sun et al. (2020a) prepared gemcitabine and polydopamine nanoparticles (PGEM/dp-16 NPs) by thin-film aqueous method for loading NLG919, a potent IDO inhibitor. In vitro experiments demonstrated that PGEM/dp-16 NP-mediated photothermal therapy (PTT) could efficiently kill cancer cells within 5 min (Figure 2B). IDO inhibitors increased more CD8⁺ T cells after releasing outside tumor cells and reduced T-reg infiltration in the TME. In addition, it can significantly reduce the tumor volume of metastatic tumor models (83%) and reduce the risk of lung metastasis (Figures 2D,E). However, in the early metastatic stage, PGEM/dp-16 NPs did not exhibit a good abscopal effect to regress distant tumors (Sun et al., 2020a). To enhance the delivery of IDO inhibitors, Li et al. (2020b) developed a mesoporous silica nanoparticle (MSN) for the delivery of doxorubicin (DOX) and IDO inhibitors. MSNs were about 70 nm in diameter and released IDO inhibitors outside tumor cells and transport DOX into tumor cells. After systemic administration to tumor-bearing mice, MSN was found to induce tumor regression and reduce the number of lung metastases. To increase the targeting of nanoparticles, Liu et al. (2019) co-encapsulated porphyrin-phospholipid conjugates and IDO inhibitors to form redox-activated liposomes. Redox-activatable liposomes prolonged the existence time of IDO inhibitors in the bloodstream, better target tumor cells. The advantage of redox-activatable liposomes was their ability to released IDO inhibitors in the reducing TME, shown the highest efficacy in inhibiting primary and distant tumors and prevented breast cancer metastasis in an orthotopic 4T1 tumor-bearing mouse model (Liu et al., 2019).

Cytokines are small-molecule proteins secreted by cells that have the functions of mediating and regulating immune processes. According to their structure and function, they are divided into interleukins, interferons, TNF, colony-stimulating factors, chemokines, growth factors, and so on. This article mainly introduces several cytokines related to the abscopal effect of killing metastatic cancer cells.

IL-12 is a potent pro-inflammatory factor produced by APCs, which induces and enhances cell-mediated immune responses. The target cells of IL-12 are NK cells and T cells, which promote the proliferation of T cells and NK cells and induce the production of γ-IFN. In addition, IL-12 increases the expression of human lymphocyte antigen (HLA) by tumor cells and promote the combination of T cells and antigens to exert anti-tumor effects (Yue et al., 1999). Although IL-12 plays an important role in anti-tumor activity, very small doses of IL-12 cause systemic toxicity (Robertson et al., 1999). The current obstacle to IL-12 treatment is the inability to effectively deliver IL-12 to tumors without causing an increase in systemic IL-12 (Nguyen et al., 2020b). Yao et al. explored a novel photothermal agent and IL-12 gene fragment co-delivery nanoparticle (CSP) (Lin et al., 2021). The hydrodynamic size of nanoparticles increased from 118 to 157 nm. Studies have found that CSP@IL-12 nanocomplex synthesis increased the level of IL-12 expression in B16F10 cells. When CSP@IL-12 nanocomplex was combined with PDT, CSP@IL-12 nanocomplex increased the expression of IL-12 gene fragments in tumors, thereby significantly increased the amount of IL-12 in the TME. CSP@IL-12 nanocomplex achieved good tumor regression, and produced the abscopal effect to kill metastatic cancer cells. In order to increase the targeting of nanoparticles to TME, Sun et al. (2020b) designed a novel nanosystem based on TAM charge-reversal polymetformin (PMet), which simultaneously delivered DOX and IL-12 gene fragments. The mechanism of increased TAM targeting lies in the synergistic encapsulation of thiolated hyaluronic acid (HA-SH) into PMet nanosystems to facilitate precise TME targeting. PMet nanoparticles with an average particle size of 192.3 nm showed improved anti-tumor and antimetastatic activity in a mouse model of breast cancer lung metastasis. However, PMet nanoparticles formed aggregates with serum components shortly after incubation. Such aggregates do not seem to be conducive to the long-term antitumor effect of PMet nanoparticles in vivo. In order to further improved the carrying efficiency, Antonio et al. used Layer-by-layer (LBL) assembly to load IL-12 gene fragments onto the surface of glyceroliposomes with poly-L-arginine (PLR) as a polymer coating for reducing systemic toxicity (Barberio et al., 2020). For the external layer, both hyaluronic acid (HA) and poly-l-glutamic acid (PLE) were chosen to produce a dense negative charge. LBL-NPs were 80–120 nm in diameter, and each LbL-NP contained approximately 50 IL-12 molecules. Reduction of IL-12 systemic toxicity and induction of abscopal effects were observed in a ovarian cancer tumor-bearing mice.

IL-2 is another cytokine of the interleukin family, also known as T cells growth factor. IL-2 binds to the IL-2 receptor and affects the proliferation, activation, and differentiation of immune cells (Raker et al., 2020). IL-2 has a certain effect on the proliferation of effector T cells, but it is not necessary for the proliferation of effector T cells. Different doses of IL-2 activate different types of immune cells (Raker et al., 2020). In the presence of low-dose IL-2, IL-2 receptors are mostly activated on Tregs; under medium- and high-dose IL-2 conditions, also lower-affinity IL-2 receptors are bound on effector T cells and NK cells (Raker et al., 2020). Therefore, it seems advantageous to maintain high-dose IL-2 levels in the tumor microenvironment. However, excessively high doses of IL-2 regulate tumor-reactive CD8⁺ T cell exhaustion by activating the aryl hydrocarbon receptor (Liu et al., 2021). The short lifespan of IL-2 has also become an obstacle to its use as an immunotherapy drug. To increase the half-life of IL-2 in the body, Votavova et al. (2015) explored combining IL-2 gene fragments with a co-polymer to make it a nanoparticle with an IL-2 agonist effect. Each IL-2 molecule carries 2 methacrylamides (HPMA). This co-polymer can prolong the half-life of IL-2 in the body and effectively control the concentration of IL-2, thereby avoiding the toxicity caused by excessive IL-2 concentration (Votavova et al., 2015). However, the biological activity of IL-2 modified by this co-polymer is weaker than that of normal IL-2. Although it causes the increase of CD8⁺ T cells and NK cells in the body, the anti-tumor effect needs to be further improved. Yin et al. co-delivered DOX and IL-2 with chitosan nanoparticles (Wu et al., 2017). Chitosan nanoparticles carrying DOX and human recombinant IL-2 solved the toxicity problem of DOX and IL-2 in the bloodstream. However, it has been found that local delivery of IL-2 increases the expression of T-regs, which was detrimental to the antitumor effect (McHugh et al., 2015; Horwitz et al., 2019). To reduce the local effect of T-regs on the antitumor effect without reducing the effect of IL-2, Jason et al. developed nanoscale liposomal polymeric gels (nLGs) for the delivery of IL-2 and TGF-β receptor-I inhibitors (Park et al., 2012). Elimination of TGF-β reverses TAM immunosuppression and enhanced antitumor immune responses. After 3 days of systemic administration, nLGs were able to maintain 38% ± 8% of initial concentrations in a mouse model of metastases. This indicates that nLGs not only increase the concentration and half-life of IL-2, but also effectively reversed the immunosuppressive environment. In another study, Sharma et al. (2020) used PEG and IL-2 to prepare a prodrug of IL-2 that could directly eliminate T-regs in TME. It selectively induced the release of cytokines on tumor-infiltrating T cells and selectively depleted Tregs in the tumor without causing changes in the number of Tregs outside the tumor (Sharma et al., 2020). Although this treatment has a good limiting effect on tumors, it has not been reported to have an abscopal effect on metastases.

Although there are not many studies on cytokine-regulated immunotherapy to trigger the abscopal effect, the effectiveness of cytokines is obvious. Nanomaterials can enhance cytokine targeting and dose sensitivity. We believe that nanomaterials to regulate cytokines to trigger the abscopal effect will have very good application prospects.

Tregs are cells that play an inhibitory role in the body. For example, Tregs produce immune tolerance by producing large amounts of TGFβ, IL10, IL35, and other factors (Schmidt et al., 2012). Tregs are produced by continuous antigen stimulation of T cells and are induced by TGF-β, and they exert their effects through TGF-β and other cytokines (Turner et al., 2020). After Tregs are activated, cytokine immunosuppression is non-antigen specific. This non-antigen specificity is not restricted by MHC and can inhibit the proliferation and activation of CD8⁺ T cells. This is the main reason for limiting distant tumor suppression (Legoux et al., 2015). The γ-IFN produced when T cells exert anti-tumor activity can regulate the development and differentiation of Tregs through interferon regulatory factor 4 (IRF4) (Alvisi et al., 2020). In the context of γ-IFN and TGF-β, peripheral CD4⁺CD25-T cells can be transformed into CD4⁺CD25 + Treg, accompanied by the up-regulation of Foxp3. CD25 is the receptor of IL-2. Tregs in tumors up-regulate the expression of the IL-2 receptor, inhibit other immune cells from binding to IL-2, and affect the activation of immune cells (DeOca et al., 2020).

An increase in the number of effector T cells will also cause an increase in the number of Tregs. Besides T cells, the function of APCs is also regulated by Tregs. The combination of Tregs and APC can cause surface molecules to peel off and inhibit the antigen presentation of APC cells (Raffin et al., 2020). DCs produce CCL22, bind to the CCR4 receptor, and regulate the migration of Tregs to the TME (Röhrle et al., 2020). Therefore, how to reduce the number of Tregs around the metastatic tumor has become a key problem in the field of tumor immunotherapy.

In addition to the aforementioned cytokine delivery that eliminates T-regs, direct depletion of T-regs in TME also enhance antitumor responses. Kohno et al. (2020) injected diphtheria toxin into tumor cells to deplete Tregs in the spleen, blood, and lymph nodes of DEREG transgenic mice after irradiation, and the survival time of the mice was prolonged. Diphtheria toxin depleted Foxp3+ Tregs in these transgenic mice. This reduces T-reg in the distal tumor microenvironment and enhances the distant effect triggered by the immune response. Depletion of Tregs induced the activation of systemic T cells and produce cytokines and cytolysins. Although depletion of Tregs throughout the body caused tumor cell regression, it also caused severe inflammation in the body (Lui et al., 2020). In one study, a homemade curcumin analog (CA) was encapsulated in α-lactalbumin (α-LA), and the Treg cell-specific antibody (mAb), as a therapeutic agent, was linked to the drug-loaded protein via matrix metalloproteinase-responded peptide (P) to form CA@α-LA-P-mAb NPs (Du et al., 2020). The diameter of CA@α-LA-P-mAb NPs were 175.0 ± 8.4 nm. It was shown that CA@α-LA-P-mAb NPs could mediate mitochondrial apoptosis, but CA@α-LA-P-mAb NPs had no toxicity to normal cells. Under the CA@α-LA-P-mAb, tumors were dramatically reduced, and the tumor inhibition rate was 75.0%. In conclusion, targeting the depletion of Tregs delayed tumor growth. In addition, increasing the ROS of TME reduced the production of T-regs. Catalase loaded onto polydopamine -ICG nanoparticles and pH-sensitive polymethacrylate-coated CaO₂ nanoparticles enhanced ROS generation in tumors, resulting in the reduction of intratumoral FOXP3+ regulatory T cells (Table 1) (Nicholas et al., 2021; Sun et al., 2021).

MDSCs are immunosuppressive cells, mainly including immature monocytes and granulocytes. Immature DCs are also a type of MDSCs. Studies have shown that ROS inhibited MDSCs differentiation (Kusmartsev and Gabrilovich, 2003). All-trans retinoic acid down-regulated the effect of ROS and regulated MDSCs differentiation by up-regulating glutathione (GSH) synthesis and down-regulating ROS content in cells (Nefedova et al., 2007). Studies have found that all-trans retinoic acid reduces MDSCs and improves the effect of immunotherapy treatment for sarcoma (Kusmartsev et al., 2003; Long et al., 2016).

The hypoxic tumor environment leads to the down-regulation of STAT3, which controls the differentiation of MDSCs to TAMs (Kumar et al., 2016b). Elimination of STAT3 expression by conditional knockout mice or selective STAT3 inhibitors can significantly reduce MDSCs expansion and T cells responses (Kortylewski et al., 2005). As an inhibitor of STAT3, docetaxel reduces tumor-induced MDSCs, TAMs, and endothelial cells infiltration into tumors, thereby inhibiting angiogenesis, tumor growth, and metastasis (Tsai et al., 2017). Interestingly, it has been clinically reported that docetaxel administered as a chemotherapeutic drug can trigger the abscopal effect to promote the regression of urothelial cancer (Ishiyama et al., 2019). PEGylated liposomes loaded with DOX nanoparticles carrying docetaxel increased the intratumoral concentration of the drug and reduce the concentration of the drug in the bloodstream, thus reducing the toxic side effects of the drug (Atrafi et al., 2020). PEGylated liposome loaded with doxorubicin was the first FDA-approved nanomedicine. It has been shown in phase one clinical trial that it increased plasma concentrations. In addition, there have been related reports on the role of TLRs and granulocyte-macrophage colony-stimulating factor (GM-CSF) in inducing MDSCs differentiation (Shirota et al., 2012; Kong et al., 2020; Long et al., 2020; Zamorina et al., 2020). These reports produced antitumor effects in primary tumor models. However, there are not many studies on the targeting of nanomaterials for MDSCs-induced immunotherapy to trigger the abscopal effect.

Many pathways affect MDSCs apoptosis. IL4Rα and STAT6 play important roles in the activation of MDSCs and the maintenance of theirs inhibitory activity by regulating the expression of ARG1 and the secretion of TGF-β (Roth et al., 2012). IL4 regulates ARG-1 transcription through the IL4Ra-STAT6 pathway and increases L-arginine depletion, reducing T cells’ survival rate (Grzywa et al., 2020). RNA aptamer specifically recognizes IL4Ra allowing it to target MDSCs, and induces MDSCs apoptosis by blocking signals from IL4Ra (Roth et al., 2012). However, IL4Ra is also expressed on B cells, T cells, and TAMs. Therefore, RNA apoptosis is not very specific to MDSCs apoptosis.

CXCR2 is the G protein-coupled receptor of human (C–X–C motif) chemokine. The expression of CXCR2 in cancer cells drives proliferation, invasion, and migration (Steele et al., 2016). However, CXCR2 is expressed on MDSCs but less expressed on tumor cells (Steele et al., 2016). Knocking out CXCR2 or CXCR2 inhibitors generate more T cells and reduce pancreatic cancer metastasis (Steele et al., 2016; Sun et al., 2019). Therefore, exploring nanoparticles to inhibit MDSCs by targeting CXCR2 inhibitors in tumors is a promising cancer treatment.

Tumor hypoxia has also been shown to recruit MDSCs. Although chemotherapeutic drugs can directly clear MDSCs, clearing MDSCs in normal tissues has the risk of inducing immunodeficiency (Zuo et al., 2020). Therefore, targeted elimination of MDSCs in the TME is a promising therapeutic modality. In a study, silica nanoparticles carrying mitochondrial respiration inhibitors improved hypoxia around tumors. The silica nanoparticles blocked the infiltration of MDSCs into the tumor and enhanced the anti-tumor effect (Tuettenberg et al., 2016; Zuo et al., 2020). He et al. developed nanoparticles (PLT NPs) using phenylboronic acid-modified low molecular weight heparin and tocopheryl succinate (Lu et al., 2021). The average particle size of PLT-NPs were 140 nm. PLT NPs could significantly improve the immune microenvironment of pancreatic tumors and inhibited spontaneous metastases. PLT NPs will not damage red blood cells as the dose increases. When such nanoparticles were injected into the body, it was observed that nanoparticles accumulated at the tumor site and reduced the infiltration of MDSCs around the tumor (by 5.02%) on the 18th day. Additionally, phenylboronic acid-modified nanoparticles reduced the number of metastases, and non-toxic heavy metal-oxo cluster secondary building units (SBUs) and photosensitizing bridging ligands enhanced the immunotherapy of metastatic cancers (Ni et al., 2019).

Although MDSCs have been demonstrated as suppressor cells in TME. However, current studies on regulating the abscopal effects of MDSCs are uncommon. This may be related to the potential of MDSCs to differentiate into other cells. The mechanism of the abscopal effect remains elusive. Therefore, it seems difficult to regulate MDSCs alone to trigger abscopal effects.

TLR7 and TLR9 are expressed on the surface of macrophages but not on the surface of tumor cells (Trinchieri and Sher, 2007). TLR agonists induce TAMs to differentiate into the M1 phenotype and trigger the abscopal effect (Shan et al., 2020; Han et al., 2021). However, due to the poor pharmacokinetics of TLR agonists, even intratumoral administration cannot achieve the best results. Zhang et al. (2020) designed a thermosensitive liposome with TLR agonists, loaded them into phosphocholine liposomes. Compared with simple intratumoral injection, thermosensitive liposomes induced more M1-type TAMs, whereas there was almost no change in M2-TAMs. The application of thermosensitive liposomes obtained longer survival rates and better long-term immune memory (Zhang et al., 2020). However, the loading rate of the thermosensitive liposomes was not enough to trigger a powerful abscopal effect. Rodell et al. explored β-cyclodextrin nanoparticles (CDNPs) for carrying TLR agonists. CDNPs were able to aggregate within tumors and were specifically captured by macrophages 24 h after injection and could more strongly differentiate M2-TAMs from M1-TAMs (Rodell et al., 2018). Unlike the high macrophage affinity of β-cyclodextrin, apoptotic bodies (ABs) as a delivery vehicle has demonstrated a strong tumor accumulation potential. Encapsulation of ABs into nanoparticles is beneficial to increase nanoparticle deposition in tumor cells. Zheng et al. (2020a) conjugated CpG to gold-silver nanorods (AuNR) and loaded them into Abs (AuNR-CpG/AB). The loading efficiency of AuNR-CpG/ABs were 3.76% ± 0.55%. More than 80% of AuNR-CpG/ABs was deposited in miscellaneous tumor cells after intravenous injection in tumor-bearing mice.

Yook et al. explored Cetuximab-targeted gold nanorods (CTX-AuNR) (Emami et al., 2021). CTX-AuNR was about 11.2 ± 1.8 nm wide and 48.2 ± 2.5 nm long. The viability of BT-20 spheroids after photoimmunotherapy (PIT) of NT-AuNR near-infrared irradiation (NIR) was much higher than that of BT-20 spheroids grown with TAM (98.6% ± 0.6%). The addition of Bi(NO₃)₃ into nanoparticles acted as a photosensitizer (PT) and enhanced the abscopal effect of PTT-triggering immune effects (Emami et al., 2021). Bi-based nanomaterials loaded with doxorubicin and hyaluronic acid induced the conversion of TAMs from M2 to M1 (Emami et al., 2021). Hyaluronic acid nanoparticles loaded with MnO₂ not only improved the local hypoxic state of the tumor but also induced the conversion of TAMs from M2 to M1 (Song et al., 2016; Parayath et al., 2018). In addition, macrophage colony-stimulating factors (M-CSFs) were key regulators of monocyte differentiation and formation and tissue-resident macrophage activity. Mao et al. used calcium carbonate as a cross-linking agent to load M-CSF into polyethylene glycol-polyglutamic acid nanoparticles (Mao et al., 2019). Nanoparticles have high drug loading efficiency (41.9% ± 12.4%). In vitro experiments showed that TAM can effectively accumulate within 1 h of injection. Nanoparticles reverse TAM phenotype and increase antitumor effect in tumor-bearing mice.

As a chemokine, chemokine (C-C motif) ligand 2 (CCL2)induces the migration of inflammatory cells and immune cells in the body. CCR2, as the receptor of CCL2, is mainly expressed in monocytes and NK cells (Hao et al., 2020). In addition, monocyte chemoattractant protein-1 (MCP-1) can also function through the CCR2 signaling pathway. CCR2 and its ligand Mcp1 pathway stimulate tumor cell proliferation (Ding et al., 2019). MCP1-CCR2 drives monocytes into the TME, which inhibits the anti-tumor activity of T cells. However, because Mcp1 inhibitors cannot be maintained in the body for a long time, the injection of Mcp1 inhibitors alone cannot cause sufficiently strong anti-tumor activity. Trac et al. (2020) explored the use of micelles to target CCR2 on monocytes and introduced apoptotic peptides into the micelles. These micelles were able to bind to monocytes in the body and induce their apoptosis, reduce the infiltration of TAMs into tumors, and promote the infiltration of T cells into the tumor, thereby enhancing the effect of immunotherapy. Shen et al. (2018) wrapped siCCR2 fragment (a siRNA for blocking CCR2 expression) in poly (ethylene glycol)-block-polylactide nanoparticles (PLG-PLA), which caused siCCR2 to target monocytes. SiCCR2 fragment-encapsulated polyethylene glycol-lactide nanoparticles. The diameter of cationic nanoparticles were 126.8 ± 15.6 nm. Cationic nanoparticles had better targeting of monocytes than neutral nanoparticles in breast cancer-bearing mice, and some of them blocked the expression of CCR2 on monocytes, blocked the infiltration of monocytes into tumor cells and reversed the immunosuppressive state in the TME. Moreover, cationic polymeric nanoparticles can promote tumors to produce CCL2 to recruit more immune cells, thereby enhancing the efficacy of chemotherapeutics (Leuschner et al., 2011; Liang et al., 2018b; Shen et al., 2018). However, these nanoparticles do not show that the abscopal effect can be enhanced.

Because of the unique biocompatibility of bismuth (Bi) and the ability of bismuth as PS to enhance PDT, Bi-based nanomaterials have been reported for bioapplications (Qin et al., 2020). Bi-based mesoporous upconversion nano-supported DOX. Lu explored a Bi-based mesoporous upconversion nanophosphor (UCNP) loaded with doxorubicin (UCNP-DOX) (Figure 3) (Qin et al., 2020). Bi(NO₃)₃ was dissolved in 20 ml of ethylene glycol and stirred until a clear solution formed. After adding DOX and stirring for 48 h, the solvent was evaporated. The diameter of UCNP-DOX was 85 nm. In vitro experiments show nanoparticles can be internalized by tumor cells and macrophages (Figure 3B). In the tumor-bearing mouse models of lung and colon cancer, after UCNP-DOX and X-ray treatment, the signal of CD68 (M1-TAM surface marker) in tumor tissue was strong, and the signal of CD206 (M2-TAM surface marker) was weak. The phenomenon showed that the level of M1 macrophages was increased, and the level of M2 macrophages was down-regulated (Figures 3C,D). The combination of nanoparticles and radiotherapy caused more M1-TAM than M2-TAM and delayed tumor growth (Figure 3E).