Tumor cell-derived exosomes regulate macrophage polarization: Emerging directions in the study of tumor genesis and development

Abstract

As an extracellular vesicle, exosomes play an important role in intercellular information transmission, delivering cargos of the parent cell, such as RNA, DNA, proteins, and lipids, activating different signaling pathways in the target cell and regulating inflammation, angiogenesis, and tumor progression. In particular, exosomes secreted by tumor cells can change the function of surrounding cells, creating a microenvironment conducive to tumor growth and metastasis. For example, after macrophages phagocytose exosomes and accept their cargos, they activate macrophage polarization-related signaling pathways and polarize macrophages into M1 or M2 types to exert antitumor or protumor functions. Currently, the study of exosomes affecting the polarization of macrophages has attracted increasing attention. Therefore, this paper reviews relevant studies in this field to better understand the mechanism of exosome-induced macrophage polarization and provide evidence for exploring novel targets for tumor therapy and new diagnostic markers in the future.

1. Introduction

Exosomes are extracellular vesicles with a diameter of approximately 30–150 nm enclosed by a bilayer lipid membrane [1]. They play a role in the transmission of information between cells [2]. Exosomes contain transmembrane and membrane anchoring proteins that can fuse with the target cell membrane, enhancing endocytosis of the target cell and promoting exosome cargos delivered directly to the cytoplasm of target cells, such as mRNA, miRNA, lncRNA, circRNA, DNA fragments, proteins and lipids [[3], [4], [5], [6]]. miRNAs, lncRNAs, and circRNAs are noncoding RNAs that are transcribed but not translated into proteins and function to alter gene expression at the transcriptional, translational and posttranslational levels [7]. Noncoding RNAs can regulate tumor glycolysis, amino acid metabolism and lipid metabolism reprogramming to influence tumor outcomes, such as promoting cancer cell growth, invasion, metastasis, angiogenesis and drug resistance [8]. These cargos can lead to the development of tumors by activating specific signaling pathways [9]. Although exosomes can be secreted by a variety of cells, exosomes secreted by tumor cells are involved in the exchange of substances and information between tumor cells and surrounding cells [10,11]. For example, upon communicating with immune cells [12], they inhibit their immune responses, promoting tumor growth and even creating a premetastatic niche conducive to tumor metastasis [13]. In addition, a hypoxic environment can increase the release of exosomes and promote tumor progression [14,15].

The tumor microenvironment (TME) is where tumor cells communicate with surrounding cells such as immune cells and the extracellular matrix [16]. As a type of immune cell, macrophages can exert immunomodulatory effects through phagocytosis, exogenous antigen presentation, and secretion of cytokines and growth factors. Notably, macrophages can be polarized into two subtypes, antitumor classically activated M1 phenotype and pro-tumor alternatively activated M2 phenotype [17]. Among them, M2 macrophages can secrete chemokines, including CCL2, CCL17 and CCL22, to promote tumor progression [18]. These two subtypes coexist in the tumor microenvironment and produce a corresponding effect when one type is predominant. The polarized phenotype can be identified by macrophage markers [19].

In recent years, with the deepening understanding of the TME, an increasing number of studies focus on the tumor microenvironment and tumor therapy [[20], [21], [22], [23]]. It has been found that the interaction between tumor cells and macrophages can have an impact on the development of primary tumors. Macrophages that take up exosomes secreted by tumor cells can be polarized into M1/M2 phenotypes (Fig. 1) [[24], [25], [26]]. This process is mostly mediated by various molecules in exosomes, which regulate target gene expression and affect tumor progression. This review summarizes the mechanism of exosome-induced polarization of macrophages and provides evidence for exploring new targets for tumor diagnosis and treatment.

Fig. 1.

Tumor cell-derived exosomes regulate macrophage polarization.

2. Signal pathways

2.1. PI3k/AKT

The PI3K/AKT pathway is involved in regulating cell survival and proliferation. Activated PI3Ks cause AKT recruitment by phosphorylating PIP2 (phosphatidylinositol-4,5-bisphosphate) to produce PIP3 (phosphatidylinositol-3,4,5-triphosphate), and activation of this signaling pathway plays an important role in tumorigenesis [27]. The following studies also suggest their involvement in the regulation of macrophage polarization.

PTEN (phosphate and tension homology deleted on chromosome ten) is a negative regulator of the PI3Kγ signaling pathway [28], which is involved in macrophage polarization [29]. PTEN mutations lead to the accumulation of PIP3 and PIP2, activating the PI3K signaling pathway [27]. PTEN downregulation can be mediated by miR-301a-3p, activating the PI3Kg/mTOR pathway [30]. miR-301a-3p expression is upregulated in pancreatic cancer cells and cell-derived exosomes in hypoxic environments, inducing macrophage M2 polarization and promoting the migration and epithelial cell mesenchymal transformation (EMT) of pancreatic cancer cells. Mice treated with exosomes derived from hypoxic pancreatic cancer cells containing miR-301a-3p had more pronounced lung metastatic nodules. Moreover, miR-301a-3p expression was reported to be negatively correlated with TNM staging and prognosis.

miR-934 is expressed in colorectal cancer (CRC) tissue, serum, cells, and cell-derived exosomes [31] and is positively correlated with colorectal cancer liver metastases (CRLMs) and adverse prognosis. miR-934 can induce macrophage M2 polarization under the regulation of the RNA-binding protein hnRNPA2B1 [32] and downregulate PTEN genes, activating the PI3K/AKT signaling pathway [33]. Polarized macrophages enhance the invasion and liver metastasis of weakly metastatic CRC cells. Liver metastases are associated with miR-934-mediated activation of the CXCL13/CXCR5 axis. The chemokine CXCL13 and its receptor CXCR5 bind to activate the NF-κB pathway to amplify the inflammatory response in tumor cells [34], promoting p65 phosphorylation. Inhibition of NF-κB/p65 signaling reduces the expression of miR-934, indicating that miR-934 first activates the PTEN/PI3K/AKT polarization pathway to promote macrophage differentiation to the protumor phenotype and then accelerates colorectal cancer metastasis via the CXCL13/CXCR5/NF-κB signaling pathway.

Activation of the PI3K/AKT pathway [35] enhances STAT3 expression and promotes macrophage polarization [36]. As a miR-21-5p target gene, increased miR-21-5p expression inhibits the level of PTEN and activates the PI3K/Akt/STAT6 signaling pathway to promote M2 polarization [37]. Then, modulating EMT-related genes activates TGF-β signaling pathways to regulate the migration and invasion of esophageal cancer. miR-21-5p has been shown to be highly expressed in esophageal cancer cells and plasma-derived exosomes [38] and is closely related to high-risk factors for esophageal cancer and macrophage polarization, affecting the progression of esophageal squamous cell carcinoma (ESCC) [39]. Coculture of exosome-carried miR-21-5p with macrophages increased the expression of M2 macrophage markers. However, inhibition of miR-21-5p expression in M2 macrophages led to increased expression of M1 macrophage markers [38]. This indicates that there is a positive feedback mechanism between M2 macrophage polarization and esophageal cancer cell EMT in tandem through tumor cell-derived exosomes. In addition, miR-21 is also closely related to bladder cancer progression [40]. It is highly expressed in exosomes secreted by bladder cancer cells, inducing macrophage M2 polarization, thereby enhancing the migration and proliferation of cancer cells [41]. Similar to esophageal cancer, miR-21 in exosomes secreted by bladder cancer cells induces M2 polarization by downregulating PTEN and enhancing PI3K/AKT-induced STAT3 signaling activity. Similar to miR-21, miR-1231-5p and miR-92b-3p [42] also activate the PI3K/AKT/STAT3/6 pathway by downregulating PTEN, inducing macrophages to polarize to immunosuppressive types. However, the exosome inhibitor GW4869 can reverse this process and inhibit bladder tumor progression.

Programmed cell death 4 (PDCD4) is involved in the PI3K γ/Akt/mTOR signaling pathway regulating cell activities [43]. As a target gene for miR-106b, PDCD4 also participates in the regulation of macrophage polarization [44]. miR-106b, which is highly expressed in CRC cell exosomes, induces M2 polarization and promotes CRC metastasis by strengthening tumor cell EMT [45]. The expression levels of p-PI3Kγ, p-AKT, and p-mTOR in miR-106b-treated macrophages were increased, while the expression levels of PDCD4 were attenuated. In accordance with in vitro experiments, miR-106b in CRC exosomes induces M2 polarization by downregulating PDCD4 expression, activating the PI3K γ/Akt/mTOR pathway [44]. EMT of CRC cells is maintained in a positive feedback manner, enhancing the migration and invasion ability of cells. As expected, miR-106b is highly expressed in CRC tissue and plasma exosomes, which is inversely connected with overall survival time and favorably correlated with tumor fraction.

eIF4A3 (Eukaryotic initiation Factor 4A-III) regulates the expression of circFARSA in non-small cell lung cancer (NSCLC) tissues, cells, and cell-derived exosomes [46]. High expression of eIF4A3 can lead to downregulation of PTEN, activation of the PI3K/AKT signaling pathway, and induction of macrophage polarization to the M2 type, thus promoting NSCLC cell migration and EMT [46].

Circ-0001142 can be delivered to macrophages by breast cancer cell-derived exosomes and activate the PI3K/AKT pathway to regulate the M2 polarization of macrophages through competitive binding with miR-361-3p [47]. Circ-0001142 also promotes the expression of the miR-361-3p target gene PIK3CB, which could be reversed by miR-361-3p overexpression. Meanwhile, PI3K/AKT is an autophagy-related pathway [47]. Circ-0001142 can inhibit autophagy, and the change in autophagy level also affects the polarization of macrophages. Polarized macrophages can enhance the proliferation, migration, invasion, and EMT of breast cancer cells and promote liver metastasis in breast cancer mice. Therefore, the circ-0001142/miR-361-3p/PIK3CB axis in breast cancer exosomes plays an important role in inducing macrophage polarization and interrupting autophagy in tumor cells [47]. Additionally, endoplasmic reticulum stress can increase the secretion of breast cancer cell exosomes [48].

2.2. STAT3

STAT (signal transducer and activator of transcription) proteins are a family of cytoplasmic transcription factors containing STAT1-6. Among them, the STAT3 signaling pathway can be activated by phosphorylation of Janus kinases (JAKs) and cytokines and is involved in cell proliferation and cancer development [49]. Additionally, it is a common pathway for the induction of macrophage polarization.

The STAT3 signaling pathway is associated with macrophage polarization to promote tumor development [50,51] and is activated by SOCS3 [52]. miR-222-3p inhibits SOCS3 expression by binding to its 3′-UTR [53]. miR-222-3p packaged in epithelial ovarian cancer (EOC) cell-derived exosomes can be delivered to macrophages, inducing M2 polarization and enhancing the migration and invasion of EOC cells. The tumor volume of mice treated with polarized macrophages increased significantly, as well as the positive staining of CD206, CD31, and LYVE-1 in tumor tissue slices, indicating that EOC cell-derived exosomes can promote tumor vascular and lymph node regeneration. Increased phosphorylated STAT3 expression and decreased SOCS3 levels in EOC-derived exosome-treated macrophages can be observed [53]. The studies above indicate that miR-222-3p induces macrophage M2 polarization by downregulating SOCS3 and activating the STAT3 signaling pathway, accelerating EOC progression. Other studies have shown that [54] the expression levels of miR-21-3p, miR-125b-5p, and miR-181d-5p in EOC cell-derived exosomes in hypoxic environments were higher than those in the normal oxygen environment. Moreover, they can promote M2 polarization, EOC cell proliferation, and migration by regulating SOCS4/5 gene expression and activating STAT3 signaling.

High expression of miR-29b-3p in oral squamous cell carcinoma (OSCC) affects the occurrence and development of this cancer [55]. Its family member miR-29a-3p is highly expressed in OSCC cell exosomes [56], promoting tumor progression by altering macrophage polarization [57]. There is a targeted regulatory relationship between miR-29-3p and SOCS1, while the STAT6 pathway associated with macrophage polarization [58] is affected by SOCS1 [59]. In miR-29a-3p exosome-treated macrophages, the expression of M2 markers and p-STAT6 increased, followed by enhanced proliferation and invasion of OSCC cells. Arguably, OSCC cell-derived exosomes downregulated SOCS1 expression by transmitting miR-23a-3p to macrophages, activating the STAT6 signaling pathway to induce macrophage M2 polarization and promote OSCC progression.

M2 macrophage markers and polarization-related signaling pathway STAT3 phosphorylation [60] are increased after macrophages obtain exosomes from hepatocyte and hepatocellular carcinoma (HCC) cells [61,62]. This process is accompanied by increased expression of CD3⁺ T-cell-inhibiting receptors. Moreover, the lytic activity and expansion ability of T cells to HCC cells is reduced. HCC-derived exosomes generate immunosuppressive activity by educating macrophages to induce T-cell failure. Previous studies have shown that miRNAs can affect tumor development by altering macrophage polarization [[63], [64], [65], [66], [67]]. Similarly, miRNAs in HCC cell-derived exosomes can be involved in the development of HCC [[68], [69], [70], [71], [72]]. For example, the expression of M2 markers, M2 polarization-related transcription Factors C/EBPβ, and p-STAT3 in miR-146-5p-treated macrophages was elevated. This may be related to exosomes that contain miR-146-5p-mediated immunosuppression. The miR-146-5p promoter can bind to the transcription factor SALL4 (spalt-like transcription Factor 4), which is regulated by STAT3 and is highly expressed in HCC [73]. Silencing SALL4 decreases the number of M2 macrophages in mouse serum and the expression level of miR-146-5p in exosomes. As a result, T-cell inhibitory receptors and tumor volume are reduced. miR-146-5p is delivered to macrophages via exosomes and binds targetively to SALL4 to activate the STAT3 pathway, leading to M2 polarization and T-cell depletion and accelerating HCC progression [73].

Phosphorylation and activation of STAT3 can often be observed in macrophages in the TME, and activating STAT3 signaling could increase the secretion of anti-inflammatory cytokines to promote tumor progression. Blocking STAT3 signaling increases the expression of proinflammatory cytokines and exerts an antitumor effect [74,75]. When the common inflammatory cytokine IL-6 signal is activated, the IL-6 receptor gp130 [76] activates JAK tyrosine kinase and STAT3 [77] to promote tumor growth. IL-6 is also expressed in macrophages [78]. Related studies have shown that breast cancer cell-derived exosomes can reduce T-cell proliferation and NK cytotoxicity, thus facilitating tumor metastasis [79]. Exosome-educated macrophages can improve the expression levels of gp130, phosphorylated STAT3, and IL6 [80]. This is due to the transfer of gp130 to macrophages with exosomes, activating the gp130-STAT3 signal and promoting the secretion of IL-6. In addition, gp130 induces a transition to a pro-tumor type of macrophage and increases their survival, which can be reversed with the use of the gp130 inhibitor SC144 [81]. Breast cancer cell-derived exosome gp130 plays a key role in the tumor microenvironment by activating the IL-6/STAT3 pathway in macrophages to tilt macrophages toward the pro-cancer phenotype. Another study found that in a hypoxic environment, IL-6 and miR-155-3p in glioma (GBM) cells can induce polarization of M2 macrophages by activating STAT3 and CAMP-responsive element-binding protein 3 (CREB3), becoming autophagy initiators [82] and reactivating the STAT3 pathway [83]. Similar to breast cancer, IL-6 and miR-155-3p are both expressed in glioma cell-derived exosomes [84] and can be delivered to macrophages through exosomes to promote the development of gliomas. The use of the STAT3 inhibitor S3I-201 inhibits IL-6 expression, blocking macrophage autophagy, suggesting that IL-6 triggers macrophage autophagy by activating the STAT3 pathway. miR-155-3p-triggered autophagy cannot be blocked by S3I-201 but instead participates in IL-6-induced autophagy by lowering the CREB3 regulator (CREBRF).

The target gene telomere repeats binding Factor 2 interacting protein (TERF2IP) of miR-1246 is involved in the regulation of the NF-κB and STAT3 signaling pathways [[85], [86], [87]]. Activating STAT3 and inhibiting NF-κb can induce polarization of M2 macrophages [88]. However, miR-1246 is expressed in exosomes derived from hypoxic glioma cells (H-GDEs). Hypoxia can affect the content of glioma-derived exosome (GDE) cargo, enhancing the expansion and immunosuppressive functions of myeloid-origin inhibitory cells (MDSCs) [89,90]. miR-1246 is also enriched in cerebrospinal fluid (CSF) in preoperative patients with malignant glioma (GBM) and significantly reduced in postoperative CSF. Elevated expression of miR-1246 in excised GBM tissue is accompanied by increased expression of M2 macrophage markers. H-GDE-treated macrophages induce M2 polarization and promote tumor cell proliferation, invasion, and metastasis. Finally, H-GDE-derived miR-1246 induces M2 polarization to promote the malignant process of GBMs.

miR-19b-3p has carcinogenic effects [91]. LncRNA-LINC00273 can promote cancer cell metastasis [92]. They have a synergistic effect in promoting the progression of lung adenocarcinoma (LUAD) [93]. Exosomes secreted by LUAD cells can induce macrophage M2 polarization to promote LUAD cell invasion and migration. This process is associated with PTPRD, a molecule upstream of the STAT3 pathway associated with M2 polarization. miR-19b-3p can bind to PTPRD (protein tyrosine phosphatase, receptor type D) and be highly expressed in exosomes. Knocking down miR-19b-3p highly increased PTPRD expression and significantly decreased the proportion of M2 macrophages. Polarized macrophages can also secrete exosomes, increase miR-19b-3p levels in LUAD cell exosomes, activate YAP signaling, and promote cell migration and invasion [93]. In addition, STAT3 can regulate the expression of LINC00273 in macrophages. LINC00273 is upregulated in miR-19b-3p-induced macrophage-derived exosomes, but expression levels in LUAD cells are not affected by miR-19b-3p. This indicates that LINC00273 is transferred to LUAD cells mainly through exosomes secreted by macrophages. After being transferred to LUAD cells, LINC00273 induces LATS2 ubiquitination by recruiting the ubiquitin ligase NEDD4, downregulating TEAD4, regulating the transcription of RBMX, activating the Hippo/YAP pathways, and promoting miR-19b-3p packaging into LUAD cell-derived exosomes. Inhibition of miR-19b-3p and LINC00273 can reduce the occurrence of LUAD [93]. Overall, miR-19b-3p in LUAD cell exosomes can induce Linc00273 transcription and M2 macrophage polarization, which in turn will be secreted by exosomes. LINC00273 is delivered to LUAD cells, facilitating the packaging of miR-19b-3p into exosomes secreted by LUAD cells and promoting the progression of LUAD [93].

Leptin can promote tumor progression by regulating the STAT3 pathway [94]. Meanwhile, leptin is highly expressed in exosomes derived from gallbladder carcinoma (GBC) cells and can be internalized by macrophages to activate the STAT3 signaling pathway and induce M2 polarization [95]. M2 macrophages can further enhance the proliferation and migration of GBC cells. Interference with leptin expression or inhibition of STAT3 phosphorylation could reverse these effects.

In addition to M2 polarization, tumor-derived exosomes can also induce macrophage M1 polarization. Protein tyrosine phosphatase receptor type O (PTPRO) inhibits breast cancer progression by dephosphorylation, leading to ERBB2 signal inactivation and endosomal internalization of ERBB2 [96]. Breast cancer cell-derived exosomes loaded with PTPRO significantly increased the proportion of M1 macrophages in the TME and reduced breast cancer cell invasion and migration [97]. PTPRO knocks down cell-derived exosomes and has the opposite effect. In addition, STAT3/STAT6 phosphorylation is inactivated by PTPRO dephosphorylation. In summary, PTPRO in tumor cell-derived exosomes can promote macrophage differentiation into an M1-like phenotype by inactivating STAT signaling to inhibit breast cancer progression.

2.3. NF-κb

NF-κB is a transcription factor involved in inflammation and carcinogenesis and belongs to the Rel family of proteins, including RelA/p65, RelB, c-Rel, p50 and p52, which are regulated and activated by IKKα and IKKβ [98]. NF-κb plays an important role in cancer pathogenesis; for example, the NFκβ1 signaling pathway is associated with colorectal carcinogenesis and with the radiosensitizing effect of ursolic acid, and miR-9/NFKB1 plays an important role in radiotherapy resistance in colorectal cancer [99,100]. Gastric cancer cell exosomes can mediate mesenchymal stem cell activation [101], induce macrophage pro-tumor polarization and enhance the proliferation and migration ability of gastric cancer cells [102]. In this process, the phosphorylation of the classical signaling element NF-κb increased, and the expression of proinflammatory factors was also elevated, indicating that gastric cancer cell-derived exosomes promoted the migration and invasion of gastric cancer cells by activating the NF-κb pathway to induce macrophage polarization [103].

2.4. MAPK

MAPK contains three signaling pathways, ERK, JNK and p38. Among them, the ERK kinase family is divided into ERK1/2, which can be activated by growth factor signaling, and when this pathway is deranged, cell proliferation increases and promotes tumorigenesis [104]. Recently, it has also been shown to be involved in macrophage polarization.

CKLF-like Marvell transmembrane domain 6 (CMTM6), which is overexpressed in a variety of tumors [105], regulates the expression of PD-L1 in tumor cells and inhibits the antitumor immune response [106]. Macrophages block the antitumor immune response [107]. In particular, M2 macrophages positively correlate with a poor prognosis for early OSCC [108]. The ERK1/2 signaling pathway is involved in the regulation of macrophage polarization [109]. Studies have shown that CMTM6 is highly expressed in OSCC tissues and is positively correlated with T staging, pathological grading, and lymph node metastasis [110]. In CMTM6 high-expression OSCC tissue, the expression of CD163 and PD-L1 was increased. This suggests that CMTM6 may be associated with M2 macrophage infiltration, PD-L1 expression, and poor prognosis. Deficiency of CMTM6 in OSCC cell-derived exosomes impairs the proliferation, migration, and invasion of OSCC cells. The level of PD-L1 expression, ERK1/2 phosphorylation, and M2 polarization ability also decreased. These results confirm that OSCC cells secrete CMTM6 in exosomes by activating the ERK1/2 signaling pathway to induce macrophage M2 polarization and participate in tumor malignant progression [110].

miR-770 and its target gene, MAP3K1, are involved in regulating the development of NSCLC [111]. miR-770 is expressed at low levels in exosomes secreted by NSCLC cells. Increasing its expression can induce apoptosis of NSCLC cells, inhibit the polarization of M2 macrophages, and reduce the migration, invasion, and EMT ability of NSCLC cells. Upon adding miR-770 agonists to exosome-treated macrophages, the levels of MAP3K1, p-JNK, and p-ERK1/2 were significantly reduced. Mice with high miR-770 expression had smaller tumor volumes and increased tumor tissue apoptosis. Hence, it can be assumed that miR-770 in exosomes derived from NSCLC inhibits macrophage M2 polarization by modulating MAP3K1 to activate the MAPK signaling pathway, inhibiting NSCLC tumorigenesis [111].

2.5. CXCR

LncRNA TUC339 is abundant in exosomes derived from liver cancer cells and can promote the proliferation of hepatocarcinoma [112]. Knocking down TUC339 can lead to increased macrophage secretion of the proinflammatory cytokines IL-1β and TNF-α, strengthened phagocytosis and decreased viability [113]. TUC339 expression is higher in M2 macrophages than in the M1 type. After repolarizing M2 to M1 macrophages, TUC339 expression decreased. Interfering with TUC339 expression partially inhibits M2 polarization. Finally, microarray analysis showed that lncRNA TUC339-induced M2 polarization is associated with cytokine receptor signaling pathways and CXCR chemokine receptor binding pathways [113].

Based on the above studies, it can be concluded that the currently known pathways for inducing macrophage polarization by tumor cell-derived exosomes include PI3k/AKT, STAT3, NF-κb, MAPK, and CXCR. Activation of these pathways can cause macrophages to exhibit either pro-cancer phenotypes or antitumor phenotypes. The phenotype varies depending on which molecules are encapsulated in exosomes. All signaling pathways are summarized in Table 1 and Fig. 2.

Table 1.

Macrophage polarization related pathways, target genes and exosome contents summarized in this review.

Author	Year	Tumor	M1/M2	Exosome cargos	Target genes	Pathways
Su et al. [136]	2016	PDAC	M2	miR-155, miR-125b	NA	NA
Wang et al. [30]	2018		M2	miR-301a-3p	PTEN	PI3K/AKT/mTOR
Linton et al. [154]	2018		M2	Arachidonic acid	NA	NA
Li et al. [113]	2018	HCC	M2	LncRNA TUC339	CXCR	CRSa
Yin et al. [60]	2019		M2	miR-146-5p	SALL4	STAT3
Wang et al. [145]	2021		M2	circ-0074854	NA	NA
Wang et al. [143]	2022		M2	LncRNA HMMR-AS1, miR-147a	ARID3A	NA
Liang et al. [114]	2019	CRC	M2	LncRNA RPPH1	TUBB3	NA
Zhao et al. [31]	2020		M2	miR-934	PTEN, CXCL13, CXCR5	PI3K/AKT, NFκB/p65
Yang et al. [44]	2021		M2	miR-106b	PDCD4	PI3Kγ/Akt/mTOR
Lu et al. [116]	2020	EC	M2	circ-0048117	TLR4	NA
Song et al. [38]	2021		M2	miR-21-5p	PTEN	PI3K/AKT/STAT6
Wu et al. [102]	2016	GC	M1	NA	NA	NFκB
Xin et al. [144]	2021		M2	LncRNA HCG18, miR-875-3p	KLF4	NA
Ham et al. [80]	2018	BC	M2	Gp130	IL-6	STAT3
Moradi et al. [141]	2020		M1	miR-130, miR-33	NA	NA
Moradi et al. [142]	2021		M1	miR-130	NA	NA
Xun et al. [124]	2021		M2	miR-138-5p	KDM6B	NA
Dong et al. [97]	2021		M1	PTPRO	NA	STAT3/STAT6
Lu et al. [47]	2022		M2	circ-0001142	PIK3CB	PI3K/AKT
Ying et al. [53]	2016	EOC	M2	miR-222-3p	SOCS3	SOCS3/STAT3
Chen et al. [137]	2017		M2	miR-940	NA	NA
Chen et al. [54]	2018		M2	miR-21-3p, miR-125b-5p, miR-181d-5p	SOCS4/5	SOCS/STAT3
Xiao et al. [138]	2020	EC	M2	miR-21	NA	NA
Qian et al. [88]	2019	Glioma	M2	miR-1246	TERF2IP	NF-κB, STAT3
Xu et al. [84]	2021		M2	miR-155-3p	IL-6, CREB3	STAT3
Tong et al. [129]	2020	HNSCC	M2	miR-9	PPARδ	NA
Cai et al. [57]	2019	OSCC	M2	miR-29a-3p	SOCS1	SOCS1/STAT6
Pang et al. [110]	2020		M2	CMTM6	NA	ERK1/2
Ono et al. [146]	2020		M2	CDC37, HSP90α, HSP90β	NA	NA
Chen et al. [46]	2021	LC	M2	circ-FARSA	PTEN	PI3K/AKT
Liu et al. [111]	2021		M2	miR-770	MAP3K1	MAPK
Chen et al. [93]	2021		M2	miR-19b-3p, LINC00273	PTPRD, NEDD4, RBMX, TEAD4	STAT3, Hippo/YAP
Cheng et al. [153]	2021	Osteosarcoma	M2	Tim-3	NA	NA
Lin et al. [41]	2019	Bladder cancer	M2	miR-21	PTEN	PI3K/AKT/STAT3
Jiang et al. [42]	2021		M2	miR-92b-3p, miR-1231-5p	PTEN	AKT/STAT3/6
Zhao et al. [95]	2021	GBC	M2	Leptin	NA	STAT3

^aCRS: Cytokine receptor signaling; NA: NONE.

Fig. 2.

Schematic diagram of macrophage polarization induced by tumor cell-derived exosomes.

3. Target genes

3.1. TUBB3 (β-III Tubulin)

LncRNA RPPH1 was obtained by next-generation sequencing from tissues in patients with CRC liver metastases, and its interaction protein TUBB3 can promote CRC progression [114]. RPPH1 expression was positively correlated with CRC TNM staging and metastasis, and the overall survival and disease-free survival of patients with highly expressed RPPH1 were shorter. RPPH1 is abundant in CRC cells, cell-secreted exosomes, and patient plasma-derived exosomes [115]. In addition, RPPH1 can induce M2 polarization of macrophages. In vitro, mice injected with exosome-treated macrophages had larger tumor volumes, more metastatic nodules, and more circulating tumor cells while shortening survival time. Therefore, CRC cell-derived exosomal lncRNA RPPH1 can enhance the migration, proliferation, and EMT of CRC cells. Interfering with TUBB3 expression can reverse the above effects.

3.2. TLR4 (Toll-like receptor 4)

CircRNA-hsa-circ-0048117 is found in exosomes secreted by ESCC cells in hypoxic environments [116]. Circ-0048117 can bind to ARID3A, promoting macrophage maturation and differentiation [117] and accelerating the malignant progression of esophageal cancer [118,119]. Treating macrophages with exosomes containing circ-0048117 induces M2 polarization. As a competitive RNA for circ-0048117, silencing miR-140 can increase the number of M2 macrophages, and inhibition of circ-0048117 counteracts this effect [116]. This process is regulated by the target gene TLR4 of miR-140. Combined with blood samples and clinicopathological data of ESCC patients and healthy populations, the plasma content of circ-0048117 in ESCC patients increased, which was positively correlated with T stage, N stage, and TNM stage. Circ-0048117 in ESCC-derived exosomes under hypoxic conditions acts as a miR-140 sponge, competing with TLR4 to bind miR-140, upregulating TLR4 expression and promoting macrophage M2 polarization [116].

3.3. KDM6B (Lysine demethylase 6B)

Recent studies have suggested that histone-modified enzyme-mediated epigenetic remodeling is associated with phenotypic regulation of tumor-associated macrophages [120]. For instance, the substrate containing histone H3 (H3K27me3/2) of triglycine and diglycine residue 27 of KDM6B may participate in positively regulating the transcription of genes required by macrophages [121]. In addition, LPS and IL4 can induce KDM6B to adjust the activation of macrophages [122,123]. miR-138-5p in breast cancer cell-derived exosomes inhibits the expression of KDM6B and induces M2 polarization in macrophages [124]. Overexpression of KDM6B can specifically stimulate the activity of promoters encoding M1-related genes and reduce the level of H3K27me3. These results revealed that breast cancer cell-derived exosomal miR-138-5p promotes macrophage M2-type polarization by inhibiting KDM6B expression, an effect related to H3K27me3 levels on M1-associated transcriptional promoters.

3.4. PPARδ (Peroxisome proliferation-activated receptor δ)

Unlike HPV-negative (HPV-) head and neck squamous cell carcinoma (HNSCC), HPV-positive (HPV+) HNSCC is more sensitive to radiation therapy [125]. The HPV + HNSCC TME contains more M1 macrophages, and the HPV- HNSCC TME contains more M2 macrophages [126]. In addition, miR-9 is closely related to HPV [127]. Exosomes can deliver miRNAs to immune cells [128]. Interestingly, miR-9 is more expressed in HPV + HNSCC cell-derived exosomes than in HPV-HNSCC [129], and exosome-treated macrophages are predominantly polarized to the M1 phenotype. Correlation analysis between the expression of M1 macrophage-specific markers and HNSCC radiosensitivity showed that M1 macrophages were positively correlated with HNSCC cell radiosensitivity. The miR-9 target gene PPARδ is involved in this process. miR-9 in exosomes secreted by HPV + HNSCC cells promotes macrophage M1 polarization by inhibiting PPARδ, increasing the sensitivity of HPV + HNSCC patients to radiotherapy.

All four genes are involved in inducing macrophage polarization, but it is still unknown which signaling pathways contribute to this effect, and further research is still needed. All target genes are summarized in Table 1.

4. Others

4.1. Non-coding RNAs

4.1.1. MiRNA

miR-155 plays an important role in hematopoiesis, inflammation, tumors, and immunity [130,131]. Knocking down miR-155 induces macrophages to trigger M2/Th2 responses [132]. miRNA-125b expression is upregulated in M1 macrophages, which improves antigen presentation and increases T-cell activity to kill tumors [133] by inhibiting the effect of interferon regulator-4 [134]. miR-155 and miR-125b are specifically delivered into pancreatic cancer cells via hyaluronic acid with polyethyleneimine and polyethylene glycol (HA-PEI/HA-PEG) nanocarriers [135]. The expression of both miRNAs in their secreted exosomes is also upregulated, thus inducing macrophage reprogramming to exert antitumor invasion and metastasis [136]. Moreover, miR-940 [137] and miRNA-21 [138] contained in EOC and EC cell-derived exosomes also induced M2 polarization. In addition, miR-130 and miR-33 have an impact on macrophage M1 polarization [139,140]. They can be transferred from exosomes derived from breast cancer cells to macrophages, enhance the phagocytic ability of macrophages, induce transformation from the M2 type to the M1 type, reduce the migration and invasion of breast cancer cells, and inhibit the development of breast cancer [141,142].

4.1.2. LncRNA

Exosomal lncRNA HMMR-AS1 mediates macrophage polarization by regulating the expression of ARID3A (AT rich interactive domain 3A) through competitive adsorption of miR-147a under the action of hypoxia-inducible factor HIF-1 to promote HCC cell proliferation, EMT and tumor growth [143]. Moreover, the hypoxic environment can increase the secretion of exosomes from HCC cells. Similarly, lncRNA HCG18 from gastric cancer cell-derived exosomes increased the expression of KLF4 (Krüppel-like Factor 4) and promoted M2 polarization by decreasing miR-875-3p in macrophages [144].

4.1.3. circRNA

hsa_circ_00074854 expression is increased in HCC cell lines and their secreted exosomes [145]. Knocking down the expression of circ_00074854 can inhibit the migration, invasion, EMT, and tumor growth of HCC cells by interacting with human antigen R (HuR) and inhibiting the polarization of M2 macrophages. In vitro experiments simultaneously confirmed that tumor growth in mice treated with exosomes at low levels of circ_00074854 was impaired. circ-0074854 in HCC cell-derived exosomes can control macrophage polarization by binding to HuR, affecting liver cancer development.

4.2. Proteins

Highly metastatic OSCC cells secrete extracellular vesicles (EVs) rich in HSP90 (heat shock protein 90) α, HSP90 β, CD326, and CD9. Double knockdown of HSP90 and HSP90β significantly reduces the survival time of highly metastatic OSCC cells [146]. CDC37 (cell division control 37) plays a key role in stabilizing the HSP90 client protein [147] and has been linked to cancer progression [148]. Knockdown of CDC37 reduces the release of proteins in EVs, such as CD9 [149]. EVs secreted by highly metastatic OSCC are rich in HSP90α and HSP90β and synergistically promote cancer cell migration [150]. Triple knockdown of CDC37/HSP90α/β reduces the survival of highly metastatic OSCC cells, and there is a compensatory response in the CDC37/HSP90 system. Double or triple knockdown decreased the levels of the three proteins in metastatic OSCC cells, and triple knockdown more effectively reduced the EMT characteristics of metastatic OSCC cells. In addition, triple knockdown also reduces the release of EV proteins, Gluc-EVs and intracellular protein stabilization [151]. Above all, triple knockdown of CDC37/HSP90α/β can diminish the release of OSCC cell EVs, reduce the EMT initiation activity of metastatic OSCC cell EVs, and inhibit M2 polarization, thereby weakening the migration and invasion function of OSCC cells. Triple knockdown also impairs EV delivery activity, which is necessary for communication between cancer cells and surrounding cells in the TME.

T-cell immunoglobulins and mucin domain-3 (Tim-3) are significantly elevated in osteosarcoma, promoting the occurrence and development of osteosarcoma [152]. Tim-3 expression in human osteosarcoma cell-derived exosomes is also elevated and induces macrophage M2 polarization [153]. M2 macrophages containing Tim-3 significantly promote osteosarcoma cell migration, invasion, and EMT. Knocking down the expression of Tim-3 in macrophages suppressed tumor growth and lung metastasis. Osteosarcoma-derived exosomes induce M2 polarization by regulating the expression of Tim-3, promoting tumor metastasis.

4.3. Lipids

Pancreatic cancer AsPC-1 cells have a high content of phospholipid esterified arachidonic acid (AA) and the highest rate of fusion with macrophages [154]. AA is a lipid that affects the fusion of exosomes with cell membranes [155]. After the removal of AA using recombinant human phospholipase A2, the fusion rate was significantly reduced, indicating that lipids in AsPC-1 cell-derived exosomes can affect exosome-macrophage fusion. After fusion, the expression of the M1 marker and immunomodulatory protein PD-L1 in macrophages did not change significantly, but the expression of the M2 marker was upregulated. Simultaneously, a variety of cytokines, chemokines, and biologically active factors (PGE2, IL-1β, VEGF, MCP-1, MMP-9, TNFα, IL-6) increased, indicating that AsPC-1-derived exosomes induce macrophages to polarize to the pro-tumor M2 type and promote the occurrence and development of pancreatic cancer.

In summary, except for breast cancer cell-derived exosomes, miR-130 and miR-33, which induce M1 polarization to exert antitumor effects, the rest of the exosome cargos induce M2 polarization to promote tumor progression. However, the specific mechanism has not been clarified, and its downstream signaling pathways can be further explored in the future. All exosome cargos are summarized in Table 1.

5. Conclusion

With the progress of continuing research, tumor treatment has gradually developed to individualization and precision in recent years. For instance, nanotechnology has been applied to the diagnosis and treatment of tumors. The use of artificial nanomaterials can more accurately transport drugs to target organs, increase drug concentrations, and reduce toxic side effects on normal tissues. However, as an exogenous material, there are still certain shortcomings, such as loss of targeting ability and rapid clearance after completion of administration. Exosomes secreted by a variety of cells in the human body are similar to an endogenous nanomaterial, which plays the role of interacting between cells, transmitting proteins, DNA, RNA, and lipids to adjacent cells. Moreover, they participate in the process of inflammation regulation, angiogenesis and tumorigenesis. Exosomes secreted by tumor cells can also change the function of surrounding cells by creating a microenvironment conducive to tumor growth and metastasis. Macrophages in the microenvironment, for example, can be polarized into two different phenotypes, antitumor growth M1 type or pro-tumor growth M2 type, by phagocytosis of tumor cells secreting exosomes. If one could determine which component in the exosome affects macrophage polarization, it might be possible to restore or enhance the antitumor function of macrophages by changing the expression of this component. To this end, this review summarizes recent studies in this field and finds that tumor cell secretory exosomes can influence tumor growth, metastasis, and drug resistance by activating multiple classical signaling pathways that induce polarization in macrophages. For example, esophageal squamous cell carcinoma can induce M2 polarization through the PI3k/AKT pathway to promote tumor angiogenesis [156], renal clear cell carcinoma can promote M2 polarization through STAT3 to enhance tumor proliferation [157], NF-κ b pathway activation induces M2 polarization to promote pancreatic cancer progression [158], M2 polarization accelerates gastric cancer liver metastasis by inducing angiogenesis and promoting intrahepatic metastatic microhabitat formation associated with ERK pathway activation [159], and MAPK pathway activation of M2 polarization can lead to temozolomide resistance in glioblastoma [160]. Currently, drugs that block activation of such pathways for antitumor purposes have been extensively studied, such as PI3K inhibitors in combination with endocrine therapy to prolong progression-free survival in hormone receptor-positive patients with advanced breast cancer [161]; mTOR inhibitors to improve the prognosis of hepatocellular carcinoma treated with transplantation, especially in patients with high postoperative AFP [162]; STAT3 inhibitors to improve overall survival in pSTAT3-positive patients with progressive colorectal cancer [163]; selective blockade of upstream and downstream targets of the NF-κB pathway, such as TNF-α, to stabilize renal cancer progression [164]; and pediatric low-grade gliomas and plexiform neurofibromas to benefit from MAPK/ERK targeted therapy [165]. There have been recent studies on targeted therapies targeting molecules carried by exosomes. For example, through engineered exosomes loaded with STAT6 antisense oligonucleotides, STAT6 was selectively silenced in M2 macrophages to reprogram them to M1, reshape the tumor microenvironment to an antitumor state, and significantly inhibit tumor growth in colorectal and hepatocellular carcinoma models [166]. This method uses exosomes to deliver targeted drugs to macrophages, altering their polarization without depleting the entire macrophage population. Taking this as an inspiration, future targeted therapies could be aimed at regulating the expression of upstream molecules related to the polarization pathway carried by exosomes, but there are still many questions to be explored. Examples include exploring more signaling pathways by which tumor cell-derived exosomes regulate macrophage polarization, extracting tumor cell-derived exosomes more precisely without the influence of secreted exosomes from other cells, determining whether tumor-derived exosomes can enter the blood circulation and affect distant organs, and determining whether existing antitumor drugs can exert antineoplasm effects by changing the way exosomes affect the polarization of macrophages. In summary, this paper aims to provide new ideas for tumor therapy by summarizing the mechanism of tumor-derived exosome-induced macrophage polarization.

Author contribution statement

All authors listed have significantly contributed to the development and the writing of this article.

Funding statement

This work was supported by National Natural Science Foundation Of China {81572408}, Natural Science Foundation of Jiangsu Province {BE2015712} and The Program of Medical Innovation Team and Leading Medical Talents in Jiangsu Province {2017ZXKJQW09}.

Data availability statement

This study is a review article.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.