Tumour-associated macrophages: versatile players in the tumour microenvironment

Tumour-Associated Macrophages (TAMs) are one of the pivotal components of the tumour microenvironment. Their roles in the cancer immunity are complicated, both pro-tumour and anti-cancer activities are reported, including not only angiogenesis, extracellular matrix remodeling, immunosuppression, drug resistance but also phagocytosis and tumour regression. Interestingly, TAMs are highly dynamic and versatile in solid tumours. They show anti-cancer or pro-tumour activities, and interplay between the tumour microenvironment and cancer stem cells and under specific conditions. In addition to the classic M1/M2 phenotypes, a number of novel dedifferentiation phenomena of TAMs are discovered due to the advanced single-cell technology, e.g., macrophage-myofibroblast transition (MMT) and macrophage-neuron transition (MNT). More importantly, emerging information demonstrated the potential of TAMs on cancer immunotherapy, suggesting by the therapeutic efficiency of the checkpoint inhibitors and chimeric antigen receptor engineered cells based on macrophages. Here, we summarized the latest discoveries of TAMs from basic and translational research and discussed their clinical relevance and therapeutic potential for solid cancers.


Introduction
Tumour microenvironment (TME) is crucial for cancer initiation, progression, and drug resistance.TME is formed by various fundamental constituents including stromal cells and immune cells (Cassetta et al., 2019;Li et al., 2023;Wang et al., 2023).Cancer development can be facilitated by tissue inflammation (Nost et al., 2021;Rajamaki et al., 2021).Despite the diverse inflammatory components in various cancer types (Cheng et al., 2021), increasing evidence demonstrated the importance of macrophages in the progression of solid cancers (Christofides et al., 2022).Macrophage is the key inflammatory effector cells, better understanding its roles may uncover effective therapeutic strategy for cancer (Coussens et al., 2013).
Clinical studies highlight the crucial roles of macrophages in cancer therapy response and resistance, including chemotherapy, radiotherapy, and PDL1-based immunotherapy (Furuse et al., 2020;Liu et al., 2020).Moreover, clinical trials of macrophage-targeted therapies have been started such as the engineered mononuclear phagocytes (Brempelis et al., 2020) and chimeric antigen receptor macrophages (CAR-M) (Klichinsky et al., 2020;Wang et al., 2022), these therapeutic approaches stem from bench-top discoveries like recruitment and differentiation (Hannan et al., 2023), functional reprogramming (Willingham et al., 2012), and integration (Dang et al., 2021), highlighting the importance of basic research and preclinical study for the development of effective cancer treatment.
In this review, we systematically summarized the functional roles and underlying mechanisms of macrophages in TME for cancer formation and progression, their translational potential, and related studies on patients for overcoming the barriers of conventional cancer treatments as well as the latest immunotherapy resistance in the clinic.Finally, we also discussed the prospects and further directions of TAMs in the clinical development for cancer treatment.
TAMs transformation also contributes to cancer progression.Besides M1/M2 polarization, single-cell RNA-sequencing revealed new TAM phenomena.Macrophage to MNT, a process where TAMs transform into neuron-like cells contributing to the formation of cancer pain (Tang et al., 2022b).MMT, where TAMs trans-differentiate into myofibroblasts for increasing abundance of pro-tumour cancer-associated fibroblasts (CAFs) in TME, enhancing the progression of non-small-cell lung carcinoma (NSCLC) (Tang et al., 2022a).

Macrophage-targeted antitumour therapy
TAMs are essential for cancer immunotherapy (Lin et al., 2019).Macrophage-targeted treatments often deplete macrophages, modify their phenotypes, or enhance antigen presentation activity of TAM (Cassetta and Pollard, 2018).Combined with chemotherapy, radiation, or immunotherapy, these techniques may increase host antitumor immunity.They have been studied in animal models and clinical studies with immunological checkpoints and other immunotherapies (Table 1).

Depletion of macrophages
TAM recruitment by CCL2 and CCR2 is critical to tumour invasion and metastasis (Xu et al., 2021b).CCL2-CCR2 signaling controls the supply of circulating inflammatory monocytes (Argyle and Kitamura, 2018) and inhibiting CCR2 keeps monocytes in bone marrow, reducing TAMs at cancer sites (Flores-Toro et al., 2020).Blocking CCL2-CCR2 axis also hinders TAM recruitment, decreasing tumour incidence and enhancing CD8 + T cells antitumour activity (Teng et al., 2017;Tu et al., 2020).Another target is CSF-1, which promotes monocyte and macrophage differentiation, proliferation, and function (Stanley and Chitu, 2014).Mouse models with CSF-1R inhibition had smaller tumors and better survival (Tan et al., 2021).Small molecule inhibitors of CSF1-R have also been shown to deplete some TAMs, enhancing tumour sensitivity to chemotherapy (O'Brien et al., 2021).

Alteration of macrophage phenotypes
TAMs change into a tumour-suppressing phenotype (Liu et al., 2021) which is a promising clinical strategy for cancer treatment.Inducing M1 macrophage phenotype through the use of selective class IIa HDAC inhibitors (Li et al., 2021a) enhances T cell responses to chemotherapy and immune checkpoint blockades (McCaw et al., 2019).The CD47/SIRP-α pathway is crucial for tumour immune escape, and blocking it enhances macrophages immune killing against tumours (Wang et al., 2020;Jia et al., 2021).Cancer immunotherapy research has also focused on anti-PD-1/PD-L1 treatment (Tomlins et al., 2023).TAMs, particularly M2 TAMs, express PD-L1 on their surface and contribute to immunosuppression by promoting T-cell apoptosis (Li et al., 2022b;Shinchi et al., 2022).In vitro-transcribed mRNA could stimulate effector molecule synthesis or cell reprogramming.mRNA in an injectable nanocarrier genetically reprogrammed TAMs into antitumour effectors.Nanoparticles formulated with mRNAs encoding the transcription factor interferon regulatory factor 5 (IRF5) and its activating kinase, inhibitor of NF-B kinase subunit-β (IKKβ), reversed the immunosuppressive TME and reprogrammed TAMs, regressing tumours in mouse cancer models (Zhang et al., 2019;Petty et al., 2021).The LILRB family, specifically LILRB2, is integral to the immune evasion strategies of cancer cells (Chen et al., 2018).LILRB2, an MHC-binding protein rich in TAMs, interacts with MHC class I molecules, which cancer cells often downregulate to dodge T cell recognition (Liu et al., 2023c).Blocking LILRB2 enhances macrophage pro-inflammatory and phagocytic activity.Its effect on macrophage activation and phagocytosis is unknown (Chen et al., 2018).MK-4830, an antibody against LILRB2, showed promising results in early trials with advanced-stage tumours (Siu et al., 2022).Responses correlated with the expression of pro-inflammatory cytokines and enhanced cytotoxic T cell-mediated anti-tumour immune response (Sharma et al., 2021).These approaches have been tested with other clinical used immunotherapies like immune checkpoints for their clinical potential with animal models and clinical trials.

Innovative strategies for TAM modulation
Recent strategies explore TAM modulation.One approach involves the engineering of T cells with chimeric antigen receptors (CAR) (Maalej et al., 2023) specifically tailored to recognize and eliminate TAMs.Research shows CAR T cells targeting macrophages are effective against various solid organ tumours, including ovarian and pancreatic cancer (Sanchez-Paulete et al., 2022).Eliminating M2like FRβ+ TAMs in the murine models of ovarian cancer, colon cancer and melanoma TME through FR-specific CAR-T cells delay tumour progression and prolong life (Rodriguez-Garcia et al., 2021).These CAR-engineered T cells show potential in redirecting immune responses against the tumour.Another method focuses on harnessing invariant natural killer T (iNKT) cells (Li et al., 2021b).These cells possess innate and adaptive immune properties, CAR-iNKT cells use iNKT TCR/CD1d and CAR recognition to deplete TAMs and tumours (Simonetta et al., 2021).Recent studies harness iNKT cells to modulate TAMs, boosting antitumour responses.Other innate T cells, including MAIT, and γδT cells, have potential clinical applications as they target and eliminate TAMs (Li et al., 2022c).In synthesis, these innovative strategies signify a shift in tumour immunotherapy (Table 2).

Prospects of macrophages in cancer
TAMs are an important immune cell type that shapes TME properties.Targeting TAMs effectively blocks the progression of various cancer types.Moreover, popularity of single-cell RNAsequencing analysis enhances the mechanistic study and preclinical research of TAMs in TME (Tang et al., 2020;Tang et al., 2021a;Chung et al., 2023).Dissecting the heterogeneity and regulatory mechanism of macrophages in cancer at single-cell resolution leads to the discovery of novel macrophage-specific therapeutics targets from the TME, for example, MMT and MNT (Xue et al., 2021;Tang et al., 2022a;Tang et al., 2022b).They are emphasizing the adaptive plasticity of macrophages.MMTs, derived from M2 TAMs with protumour activities, lead to the formation of CAFs.These CAFs are key in driving cancer progression (Chen and Song, 2019;Li et al., 2020).The roles of MMT-derived CAFs in functions, including adaptive immunity suppression, drug resistance, metastasis, and promoting cancer cell stemness warrant investigation.Conversely, MNTs highlight the transformation of TAMs into neuron-like entities, influencing de novo neurogenesis in the TME (Tang et al., 2022b) and contributing to cancer-associated pain (Shepherd et al., 2018).This transition, while prevalent in NSCLC, is also seen in other tumours, emphasizing its importance in cancer pain and tumour innervation (Tang et al., 2022b).Given the impact of cancer pain on quality of life, especially in patients with advanced stages of the disease (Wang et al., 2021c), understanding MNT is vital for pain management strategies.Notably, these transitions were found to be mediated by a Smad3-centric gene network in TAMs, highlighting the potential of macrophage-targeted Smad3 interventions as a promising therapeutic approach in cancer immunotherapy (Tang et al., 2017;Feng et al., 2018;Tang et al., 2021b;Tang et al., 2022b).These new findings lead to the development of effective therapeutic approaches to enhance the efficiency of conventional anticancer treatments as well as the latest immunotherapies which are not primary or secondary resistant in patients with solid cancers (Kim et al., 2019b;Kim et al., 2020;Tang et al., 2020;Chung et al., 2021;  Xue et al., 2021).Besides, macrophages are considered as a primary target of anti-inflammatory therapy for cancer prevention, their therapeutic potential is explored by new trials worldwide (Tang et al., 2019;Lee et al., 2021;Tang et al., 2022d).Despite the challenges, a better understanding of the immunodynamics of TAM shows a substantial potential for improving the therapeutic efficiency and clinical outcomes of cancer patients in the future.

FIGURE 1
FIGURE 1 TAMs play a complex dual role in the progression of cancer.M1 TAMs contribute to the anticancer response via multiple mechanisms.They can produce reactive oxygen species (ROS) and reactive nitrogen species (RNS) to cause oxidative damage and kill cancer cells.The secretion of proinflammatory cytokines and chemokines (e.g., TNF-α, IL1B, IL12A/B, CCL5, and CXCL10) can mobilize other anticancer immune cells, like T cells and NK cells, into the TME.Anti-angiogenesis is promoted by secretion of thrombospondin-1 and angiostatic chemokines like CXCL9, CXCL10, and CXCL11.TAMs also express MHC class I and II molecules for antigen presentation to further priming and activation of T cells.The interaction between CD80/ CD86 on TAMs and CD28 on T cells provides a second signal for T cell activation.M2 TAMs promote immunosuppression, angiogenesis, and tumour growth/metastasis while contributing to drug resistance.Immunosuppression involves secretion of TGF-β and IL-10, expression of PD-L1, and CCL22induced Treg activation.In angiogenesis, TAMs secrete factors like VEGF, FGFs, PDGF, HGF, MMPs, and IL-8/1.During tumour growth and metastasis, M2 TAMs enhance proliferation, migration, and invasion.Factors like EGF, PDGF, VEGF, CCL-10, and MMPs play key roles.TAM can also undergo transformation to MNT and MMT, resulting in the generation of cancer pain and cancer-associated fibroblast.In drug resistance, TAM-derived TGF-β, IL-6/8, and PDGF stimulate survival pathways and enhance DNA repair in cancer cells.It is noteworthy that macrophages can switch from M1 phenotype to M2 phenotype during tissue repair.

TABLE 1
Selected clinical trials of drugs targeting TAMs.

TABLE 1 (
Continued) Selected clinical trials of drugs targeting TAMs.

TABLE 1 (
Continued) Selected clinical trials of drugs targeting TAMs.
(Continued on following page)Frontiers in Cell and Developmental Biology frontiersin.org

TABLE 1 (
Continued) Selected clinical trials of drugs targeting TAMs.
(Continued on following page)Frontiers in Cell and Developmental Biology frontiersin.org

TABLE 1 (
Continued) Selected clinical trials of drugs targeting TAMs.

TABLE 2
Innovative strategies targeting TAMs in tumour microenvironment.FG tumour cells, have dual CAR/TCR targeting mechanisms, sustain antitumour capacity in presence of macrophages, and target TAMs