The hepatic macrophage: a key regulator of liver metastatic tumor microenvironment through cell crosstalk

Abstract

Liver metastasis continues to be a leading cause of cancer-related mortality, particularly in colorectal, pancreatic, and breast cancer. The successful establishment of metastatic lesions depends critically on the liver metastatic tumor microenvironment, where reciprocal cellular interactions between disseminated tumor cells and both parenchymal hepatocytes and non-parenchymal cells facilitate tumor cell colonization and outgrowth. Hepatic macrophages that encompass both tissue-resident Kupffer cells (KCs) and monocyte-derived macrophages (Mo-Macs) have emerged as pivotal regulators of liver metastatic progression. This review summarizes recent progress from the following perspectives: (I) Primary tumors recruit macrophages via secretion of cytokines and exosomes, or induce phenotypic alterations in resident KCs and recruited Mo-Macs, thereby establishing a premetastatic niche; (II) Once the premetastatic niche is formed, hepatic macrophages directly interact with tumor cells to mediate their capture, colonization, and subsequent outgrowth; (III) Furthermore, hepatic macrophages regulate phenotypic changes in T cells, NK cells, hepatocytes, and hepatic stellate cells (HSCs) through cytokine/exosome secretion or direct cell-cell interactions, which induce T cell exhaustion, impairment of NK cell cytotoxicity, and activation of HSCs leading to fibrotic microenvironment formation. Additionally, we review advances in macrophage-targeted therapeutic strategies against liver metastasis. By delineating the pivotal roles of hepatic macrophages in metastatic progression and analyzing current clinical limitations of targeting macrophages for liver metastasis therapies, this review provides foundational insights for understanding macrophage biology and developing effective therapeutics.

Background

Distant metastasis is the leading cause of cancer-related mortality [1, 2], and the liver is one of the most common and prognostically unfavorable sites of cancer metastasis [3, 4]. Clinical observations reveal that 50% of cancer patients across various malignancies develop liver metastases during disease progression [5]. Liver metastasis predominantly originate from tumors of colorectal cancer (CRC), followed by pancreatic cancer, breast cancer, melanoma and lung cancer [6]. Notably, compared to primary liver cancers, liver metastasis occurs 18–40 times more frequently than primary hepatic malignancies, with liver metastatic patients demonstrating significantly poorer prognoses [7, 8].

Liver metastasis develops through a complex multistep biological process comprising three main steps: pre-metastatic niche establishment, tumor cell colonization, and metastatic lesion formation. Inherently, several metastatic-friendly features of the liver are essential for cancer liver metastasis. First of all, the liver possesses a dual blood supply system: the hepatic artery provides oxygenated blood, while the portal vein delivers nutrient and toxins to liver [3, 9]. Therefore, the liver has an abundant blood supply, which creates an environment conducive to tumor cell colonization. Structurally, the hepatic lobules are composed of hepatic sinusoids that decelerate the speed of blood flow [3, 9]. This characteristic facilitates tumor cell adhesion and subsequent extravasation into hepatic tissues. Furthermore, the liver, as an important organ regulating the energy balance, offers metastatic tumor cells access to nutrients and energy substrates [10]. Significantly, the liver is recognized as a crucial immunotolerant organ. Due to the continuous influx of gastrointestinal-derived toxins and food antigens via the portal vein, the liver undergoes persistent antigenic stimulation, which drives the adaptation of immunosuppressive inclination in the liver. The immunotolerant microenvironment of the liver is collectively maintained through synergistic interactions among hepatic resident cells, such as Kupffer cells (KCs), liver sinusoidal endothelial cells (LSECs), and hepatic stellate cells (HSCs). KCs, positioning on the luminal side of the sinusoids, perform dual immunological functions by phagocyting debris and pathogens through specialized scavenger receptors, and modulating immune homeostasis via interplay with other cells. LSECs, as the main component of sinusoidal endothelium, can act as antigen-presenting cells to activate T cells. Additionally, residing in the perisinusoidal space of Disse, HSCs dynamically regulate immune responses through interacting with KCs as well as recruiting immune cells to maintain chronic inflammation and fibrogenesis [10, 11].

As to well known, tumor microenvironment (TME) is essential for tumor progression and metastasis. In liver metastasis, the metastatic TME contributes to metastatic tumor cell colonization and proliferation, as well as immunosuppression [4]. Strikingly, macrophages represent an essential component of the liver and play a crucial role in the liver metastatic TME [12–14]. Hepatic macrophages are mainly composed of resident KCs and infiltrating macrophages. Recently, there are numerous studies aim to clarify the role of hepatic macrophages in liver metastasis [13]. On one hand, in the primary tumor stage, tumor-derived factors, such as cytokines and exosomes, can recruit macrophages into the liver, which helps to create a pre-metastatic niche for tumor cells [15, 16]. On the other hand, the resident KCs are the first line for defending the circulating tumor cells (CTCs) colonization through arresting and phagocytosing tumor cells [17, 18].

Cell-to-cell communication is a vital mechanism by which macrophages regulate the metastatic TME of the liver [19, 20]. Macrophages interact with metastatic tumor cells directly and promote proliferation and colonization of metastatic tumor cells, but also have an indirect effect on non-tumor cells, such as immune cells, hepatocytes, and (HSCs, to mediate the formation of liver metastasis [19]. In the present review, we describe the distinct ontogeny and population of hepatic macrophages, the crosstalk between metastatic tumor cells and hepatic macrophages, the interaction of hepatic macrophages and non-tumor cells such as immune cells and hepatocytes in the liver, and the latest insights into therapeutic strategies by targeting hepatic macrophages in liver metastasis.

The ontogeny, composition and function of hepatic macrophage in the liver

Macrophages are pivotal phagocytic cells within the immune system, derived from monocytes and widely distributed across various tissues [21]. They engage in both innate and adaptive immune responses by phagocytosing pathogens, dead cells, and foreign particles, thereby performing essential functions in host defense, tissue repair, and immunoregulation [21]. Depending on their tissue localization, macrophages differentiate into specific subtypes, such as alveolar macrophages in the lungs and Kupffer cells (KCs) resident in the liver. Additionally, they can polarize into M1 or M2 phenotypes in response to inflammatory or reparative contexts [21]. In the liver, hepatic macrophages are mainly divided into two distinct subsets of resident KCs and monocyte-derived macrophages (Mo-Macs) [22].

It is widely accepted that resident KCs originate from fetal liver-derived erythromyeloid progenitors [23]. In mice, yolk sac-derived erythromyeloid progenitors directly set seeds into the fetal liver to give rise to KCs or develop into circulating monocytes which subsequently migrate to the fetal liver in a CX3C chemokine receptor 1 (Cx3cr1) dependent way and differentiate into KCs [24, 25]. Mouse KCs express several cell markers such as F4/80, Vsig4, Clec4f, and Tim4. In mice, KCs can be distinguished from Mo-Macs by F4/80^high, Cd11b^int, Clec4f⁺, and Cx3cr1⁻. KCs are the main hepatic macrophage population, accounting for approximately 90% of liver macrophages during liver homeostasis [26]; however, the proportion of KCs decrease in liver disease (Fig. 1). For example, using Membrane-spanning 4-domains subfamily A member 3 (Ms4a3) as a lineage tracing marker for Mo-Macs in the liver in which KCs were Ms4a3⁻ and Mo-Macs were Ms4a3⁺ showed that the proportion of KCs was around to 90% in the healthy liver, but this proportion decreased to 70% in fibrotic liver induced by carbon tetrachloride [26]. KCs are located in the liver sinusoid where they interact with parenchymal and non-parenchymal cells such as hepatocytes and HSCs, which are important for maintaining the self-renewal capability and proper functions of KCs. KCs play a pivotal role in maintaining liver homeostasis owing to their phagocytic functions by which they clear cellular debris, metabolic waste, and pathogenic microorganisms, and phagocytose red blood cells to maintain iron homeostasis [27–29]. In addition, KCs can regulate cholesterol homeostasis by producing cholesteryl ester transfer proteins [30].

Fig. 1.

The composition and ontogeny of hepatic macrophages. Hepatic macrophages consist of resident Kupffer cells (KCs) as well as recruited Mo-Macs. (A) In homeostasis, KCs originate from yolk sac-derived erythromyeloid progenitors and highly express surface markers such as F4/80, Clec4f, and Tim4, while exhibiting low expression of Ly6c. Recruited macrophages are primarily differentiated from bone marrow-derived monocytes and show high expression of Ly6c and Ccr2. In addition, a small population of Mo-Macs is derived from the spleen and exhibits low expression of both Ly6c and Ccr2. (B) Under disease conditions, a mass of Mo-Macs are recruited into liver to mediate inflammation response, while the percent of KCs are decreased

Although murine hepatic macrophage ontogeny and developmental pathways have been well characterized, the embryonic origins of their human liver macrophage remain poorly understood, possibly due to both limited accessibility to developmental-stage human specimens and the lack of sufficiently precise methodologies to identify embryo-derived hepatic macrophages [31]. Recently, a study using single cell sequencing (scRNA-seq) to characterize macrophages in human embryos has revealed evolutionarily conserved origins similar to murine models; specifically, one subset of macrophage populations originates from in situ differentiation at the early stage of yolk sac and subsequently disseminates into developing embryonic tissues. In contrast, another distinct subset derives from yolk sac-derived primitive myeloid progenitors that undergo differentiation into transitory monocytes in the fetal liver before maturing into definitive macrophage populations [31]. Furthermore, in contrast to murine KCs that have well-defined markers, there is currently no consensus for identifying human KCs markers. For instance, Clec4f serves as a classical marker distinguishing KCs from other hepatic macrophages in mice, while Clec4f is not conserved in human KCs [32]. Although scRNA-seq was applied to shed light on the true identity of human KCs, each study reached different conclusions regarding human KC markers [33, 34]. Recent research combining scRNA-seq and spatial transcriptomics has systematically characterized conserved markers of tissue-resident KCs across seven species including human and mouse. This study proposes that CD5L, VSIG4, CD163, Folate receptor 2 (FOLR2), macrophage receptor with collagenous structure (MARCO), and Solute carrier family 40 member 1‌ (SLC40A1) can constitute core genetic signatures for defining human KCs, providing a potential framework for standardized identification across species [23, 25, 32] (Table 1).

Table 1.

Protein and genes of KCs in murine and human liver

Marker	Mouse	Human	Conserved
F4/80	M
VSIG4			C
FOLR2			C
CD163			C
CLEC4F	M
CLEC2	M
TIM4			C
CD5L			C
MARCO			C
GFRA2			C
ADRB1			C
TMEM26			C
SLC40A1			C
HMOX1			C
SLC16A9			C
VCAM1			C
SUCNR1		H
CD207			C

M: mouse; H: human; C: conserved

The hepatic macrophage pool can be rapidly replenished by Mo-Macs, which is mainly derived from monocytes. In mice, bone marrow resident hematopoietic stem cells can differentiate into monocytes that enter circulation and develop into Mo-Macs in the liver under specific conditions, especially during liver diseases [24, 35]. There are two major groups of Mo-Macs in the liver: Ly6c^high, which are characterized as CCR2⁺CX3CR1⁺CD43⁻ cells, and Ly6c^low monocytes, which are defined as CX3CR1⁺CCR2⁻CD43⁺ cells [21] (Fig. 1). Importantly, it has been reported that Ly6c^high monocytes originate from the bone marrow, whereas the spleen is an important source of Ly6c^low monocytes [36, 37]. During homeostasis, only 10% of hepatic macrophages are Mo-Macs, but this proportion can rapidly increase when the liver is injured, such as in ischemia reperfusion, liver fibrosis, cirrhosis, and liver cancer [25, 38, 39]. Mo-Macs play an essential role in regulating iron and cholesterol metabolism in the healthy liver, and is important for mediating inflammation during liver diseases as well as liver cancer [40].

The biological process of liver metastasis and how primary tumors regulate the hepatic macrophage recruitment and phenotype

Liver metastasis (LM), in which primary tumor cells set seeds into the liver and finally develop into hepatic metastatic lesions, is a complicated process that requires cancer cells to interact with different cells, including parenchymal and non-parenchymal hepatic cells, and immune cells. The process of LM has been divided into several phases: (I) the initial phase, in which tumor cells undergo epithelial-mesenchymal transition and invade the surrounding tissue to acquire an invasive phenotype; (II) the pre-metastatic niche phase, in which the primary tumor can induce a survivable metastatic microenvironment for metastatic tumor cells in the liver; (III) the tumor cell extravasation and seeding phase, in which tumor cells extravasate from the circulation and seed in the liver; and (IV) the colonization phase, in which tumor cells regain the capability of proliferation and grow into metastatic lesions [41, 42] (Fig. 2).

Fig. 2.

How primary tumors regulate the hepatic macrophage recruitment and phenotype. Primary tumors can not only promote the recruitment of macrophages but also regulate their polarization during liver metastasis. (A) Recruitment of macrophages by primary tumors. Signals derived from primary tumors can be divided into five categories: cytokines, secretory proteins, exosomes, metabolites, and other mediators. In the primary tumor phase, tumor-derived CCL2, Cxcl2, and MSP recruit Mo-Macs into the liver, while uPA and IL-1α enhance macrophage adhesion. (B) Polarization of macrophages during liver metastasis. Several mechanisms induce M2 polarization of hepatic macrophages during metastasis, including receptor–ligand interactions mediated by Ca5–Ca5R, CTHRC1–TGFBR2, VEGF–VEGFR, fatty acid–CD36, IL-17D-CD93 as well as signaling via IL-18. Tumor-derived exosomes containing microRNAs (e.g., miR-151a-3p, miR-934, miR-519a-3p) and proteins such as ANGPTL1 also promote M2 polarization. Additionally, although tumor-derived CXCL16 has the potential to polarize macrophages toward an M1 phenotype, its expression is generally downregulated during liver metastasis

Establishment of a pre-metastatic niche that creates a supportive microenvironment for tumor cell colonization and proliferation is an important step in liver metastasis [43]. Hepatic macrophages are a major component of the liver microenvironment, consisting of various immune cells and liver non-parenchymal cells, in which macrophages not only help to create an immunosuppressive microenvironment but also promote the survival and growth of tumor cells [44]. Importantly, the effects of primary tumors on the liver microenvironment and hepatic macrophages are mainly dependent on the primary tumor-derived signals and their delivery to the liver by which primary tumor cells can promote the recruitment of Mo-macs into liver and regulate the phenotype transformation of Mo-macs and resident KCs [20]. Some studies have been conducted to clarify the mechanism by which primary tumors affect the liver microenvironment and hepatic macrophages. According to current studies, primary tumor signals can be classified into the follow types: (I) cytokines or secretory proteins [15, 16, 45], (II) exosomes or extracellular vesicles [46–48], (III) tumor-derived metabolites [49, 50], (IV) complement systems [51] and (V) Other media such as intestinal bacteria [52] (Fig. 2).

The secretion of cytokines/chemokines and secretory proteins is the main mechanism that induces macrophage infiltration into the liver. C-C motif chemokine ligand 2‌ (CCL2) is one of the most important chemokines responsible for monocyte recruitment by interacting with its receptor, CCR2 (Fig. 2). In CRC liver metastasis (CRLM), increased CCR2⁺ macrophage infiltration was confirmed at the sites of liver metastasis and correlated with poor prognosis, and CCL2 secreted by CRC tumor cells was responsible for macrophage recruitment [15]. Furthermore, a recent study has shown that the density of macrophages recruited by tumor-derived CCL2 increased three days after tail vein injection in a liver metastasis mouse model. Additionally, the secretion of CCL2 is regulated by Transcription Factor 4 (TCF4), which was confirmed to be highly expressed in liver metastases and positively correlated with macrophage infiltration. [45]. Notch-dependent inflammatory signaling also upregulates the expression of CCL2 in pancreatic cancer cells, which activates the ability of macrophage extravasation to promote tumor-associated macrophage infiltration at the site of liver metastasis. Importantly, pancreatic cancer cells secreted Jagged1, Interleukin-1α (IL-1α) and urokinase-type plasminogen activator (uPA) increase the ability of macrophage adhesion to enhance hepatic macrophage infiltration and M2 polarization, which were blocked with neutralizing antibodies against murine Jagged1, IL-1α, uPA receptor (uPAR) after 7 days of liver metastasis model construction [16]. In addition, secretory macrophage-stimulating protein (MSP) also recruits macrophages into the liver and enhances liver metastasis [53]. Although cytokines and secretory proteins play a major role in regulating hepatic macrophage recruitment during liver metastasis, some mediators such as the complement system and intestinal bacteria, also have a role in this process [51, 52]. The complement system has also been demonstrated to involve in recruiting macrophages to the liver during CRLM. In a mouse model of splenic injection-induced CRLM, elevated plasma complement 5a (C5a) levels were observed at 10 days post-injection. The increased C5a promoted the infiltration of macrophages, neutrophils, and dendritic cells through its interaction with the C5a receptor (C5aR). [51]. Furthermore, a recent study demonstrated that Escherichia coli, a primary tumor resident bacterium in CRC, disrupts the gut vascular barrier regulated by plasmalemma vesicle-associated protein-1 (PV-1) and disseminates into the liver through the gut-liver axis Subsequently, disseminated Escherichia coli promoted the infiltration of macrophages and inflammatory monocytes, two main components of the pre-metastatic niche, to help construct a mature pre-metastatic niche for metastatic CRC tumor cells [52].

Hepatic macrophages are highly plastic and can adapt to different metastatic microenvironments. Before tumor cell seeding in the liver, the primary tumor not only regulates the recruitment of macrophages, but can also regulate the phenotype of hepatic macrophages into metastasis-associated macrophages to promote liver metastasis (Fig. 2). Notably, compared to primary tumors, liver metastases upregulate genes encoding cytokines and secreted proteins to directly modulate macrophage phenotypes. Alternatively, aberrant expression of pathogenic genes in metastatic tumor cells facilitate cytokine secretion and induce macrophage phenotypic transformations. For example, Interleukin-6 (IL-6) highly express in metastatic CRC cell lines, compared to non-metastatic CRC cell lines, and the secreted IL-6 induced immortalized mouse Kupffer cell line into M2 polarization [54]. Furthermore, Collagen Triple Helix Repeat Containing 1 (CTHRC1), a colorectal cancer (CRC) metastasis-associated gene, demonstrates elevated expression in CRC patients developing hepatic metastases. Subsequently, CTHRC1 originating from primary tumors facilitates M2 polarization of hepatic-infiltrating macrophages through interacting with Transforming growth factor beta receptor 2 (TGFBR2) and activating downstream signaling [55]. Additionally, Vascular endothelial growth factor (VEGF) secreted by primary CRC cells upregulates Caspase recruitment domain-containing protein 9 (CARD9) expression - a critical innate immunity regulator - in hepatic macrophages via Skr activation, simultaneously driving their polarization toward metastasis-promoting phenotypes [56]. Beyond the above-mentioned, aberrant expression of N-acetyltransferase 10 (NAT10) in gastric cancer liver metastasis leads to the upregulation of CXCL2 and subsequently indue M2 polarization of hepatic macrophage with increasing CD163⁺CD206⁺ macrophages infiltration [44], and the SLIT and NTRK-like protein 4‌‌ (SLITRK4) upregulation also promote M2 polarization of hepatic macrophage by secreting CCL2, Growth/differentiation factor 15 (GDF15) and Colony-Stimulating Factor 1 (CSF-1) [57]. Moreover, WNT11 overexpress in the CRLM and PDAC liver metastasis and increase the secretion of interleukin-17D (IL-17D) to subsequently induce Cd163 and Arg1 expression in macrophages, terming as M2 polarization, through interacting with membrane receptor CD93 [58]. In addition, tumor-derived metabolites have been reported to polarize hepatic macrophages to the M2 phenotype. For instance, tumor cell-derived long-chain fatty acids are engulfed by hepatic macrophages through CD36, leading to lipid deposition and M2 polarization of macrophages in a liver metastasis mouse model [49]. While tumor-secreted factors predominantly drive immunosuppressive phenotype of hepatic macrophages, dysregulation of inhibitory cytokines that maintain macrophage-mediated anti-tumor effector functions critically contributes to metastatic progression in the liver microenvironment. Kee et al. engineered C-X-C motif chemokine ligand 16‌ (CXCL16_-overexpressing SL4 murine colorectal cancer cells and demonstrated that tumor-derived CXCL16 upregulates tumor necrosis factor α (TNFα) production in hepatic macrophages. Critically, this macrophage-secreted TNFα suppressed liver metastatic progression by inducing tumor cell apoptosis [59]. This inhibitory effect mediated by CXCL16 and TNFα indicates that restoring the levels of CXCL16 and TNFα in liver metastases may represent a promising therapeutic strategy.

Exosomes play important roles in cell-to-cell communication. Primary tumor-derived exosomes have been reported to promote liver metastasis. First, primary tumor-derived exosomes packaged with microRNAs and proteins were absorbed by resident KCs and induced a phenotype transformation of KCs, which promoted pre-metastatic niche formation [47, 60–64] (Fig. 2). Recent studies showed that primary CRC and Gastric tumor-derived exosomes carried miR-519a-3p, miR-934 and miR-151a-3p to induce M2 polarization of resident KCs and Transforming growth factor beta (TGF-β) secretion by KCs [46–48]. In addition, KCs angiopoietin-like protein 1 (ANGPTL1) has been proven to be a tumor metastasis suppressor in several cancers. However, ANGPTL1 was downregulated in CRC-derived exosomes, which promoted liver metastasis by regulating KCs secreting pattern [65].

How hepatic macrophages affect liver metastasis

Liver metastasis requires primary tumor cells to disseminate to the liver and survive in transit. During dissemination, primary tumor cells invade the basement membrane and intravasate blood vessels as CTCs. In circulation, CTCs can be coated with platelets, neutrophils, or tumor stromal cells, which protect CTCs from extensive attrition of immune stressors and redox reactions, and survive in transit [66–68]. After reaching the liver, disseminated tumor cells (DTCs) interact with different cells in the metastatic niche, particularly macrophages, to facilitate formation of metastatic lesions by promoting DTC colonization and outgrowth. In the metastatic niche, macrophages regulate liver metastasis progression by interacting directly with DTCs and other non-tumor cells in metastatic niches, such as hepatocytes, HSCs, and immune cells [69].

The direct crosstalk between macrophage and disseminated tumor cells in liver metastasis

Crosstalk between macrophages and DTCs in the metastatic niche is crucial for the survival of DTCs and subsequent clonal proliferation. However, emerging evidence suggests distinct functional differences between resident KCs and Mo-Macs in modulating liver metastasis [58, 70]. KCs, located in the liver sinusoid, commonly serve as the first line of defense against particulates, pathogens, and abnormal cells entering the portal or arterial circulation [71]. Current consensus indicates that KCs predominantly exert antitumor effects in the early stage of liver metastasis [72–75], though paradoxical pro-tumoral functions have been observed in advanced disease [74, 76] (Fig. 3). Decades ago, Bayón et al. reported that KCs arrest and clear the circulating tumor cells, infused through the mesenteric vein of rats, in the early stage of the liver metastasis model in which tumor cells can be seeded in the liver after two days of infusion [18]. Moreover, macrophages were surrounded in the small tumor cell cluster, and depletion of KCs resulted in the degradation of liver metastasis in 14 days of infusion [18], indicating that KCs play a key role in preventing DTC form seeding in the liver. Consistently, the latest research, using intravital microscopy, proved that KCs can colocalize and phagocytize DTCs to inhibit the formation of liver metastasis in the early stage, but fail to inhibit liver metastasis in the late stage due to KCs loss preferentially infiltrating the tumor core and periphery [70]. Furthermore, resident KCs directly uptake and clear DTCs via Dectin-2 and Fc receptor-mediated phagocytosis [73, 77]. In summary, the prevailing view in current studies is that KCs play an essential role in the early stage of DTCs seeding, primarily through their phagocytic function to prevent DTCs colonization.

Fig. 3.

The effects of hepatic macrophages on disseminated tumor cells and non-tumor cells in the liver. The profile of hepatic macrophages with other liver components in liver metastatic microenvironment. (A) The GATA6⁺ and SPP1⁺ macrophages can induce T cells exhaustion, apoptosis and function inhibition by GDF15/TGFBR2, PD-L1/PD-1 and Fas/FasL signaling pathways as well as increase the infiltration of Treg cells by Fn14. (B) Macrophage can secret IL-8 and Il-1β to activate NK cells and result in an anti-tumor effect. In addition, both Mo-Macs and KCs promote the activation of HSCs and promote liver metastasis, and Mo-Macs can induce HSCs activation by secreting PGRN and granulin. The crosstalk between hepatocytes and hepatic macrophages in liver metastasis is mediated by SAA/FPR1 and SAA/TLR2 signals. (C) Mo-Macs can promote disseminated tumor cell survival by secreting IL-3, IL-4, IL-18, TGF-β and CXCL13, while KCs often have an inhibitory effect on tumor cells through arresting and dectin-2/Fc receptor mediated phagocytosis

Although KCs are believed to play a double-faced role in regulating liver metastasis depending on the stage of liver metastasis, Mo-Macs primarily plays a tumor-promoting role in liver metastasis. As previously described, the primary tumor contributes to the recruitment of Mo-Macs into the liver, and Mo-macs polarize into a tumor-promoting phenotype during liver metastasis [15]. Spatial analysis revealed C-C chemokine receptors 1 (CCR1)⁺ and CCR2⁺ macrophages were shown to be enriched in periphery of the metastatic lesion area [15, 78] and accelerated the formation of liver metastasis by paracrine secretion of pro-tumoral factors, such as IL-3, IL-4, IL-18, TGF-β, and C-X-C motif chemokine 13 (CXCL13) [46, 56] (Fig. 3).

The indirect effect of macrophage on liver metastasis by interacting with non-tumor cells

Hepatic macrophages not only regulate liver metastasis by directly interacting with tumor cells but can also affect liver metastasis by altering the metastatic microenvironment through the interaction between macrophages and non-tumor cells, such as T cells, B cells, natural killer (NK) cells, HSCs, and hepatocytes [19] (Fig. 3).

Macrophages and immune cells in liver metastasis

T cells

T cells, the main components of adaptive immunity, play an important role in cancer. The extent of T cell infiltration at the tumor site has long been recognized as an important prognostic factor for cancer patients. An increasing infiltration of antigen-experienced CD8 and CD4 T cells always indicates satisfactory tumor immunity and a good prognosis, while infiltration of increasing suppressive T cells, such as regulatory T cells (Treg cells), leads to a poor prognosis [79]. Previous studies reported that a low density of CD8⁺ T cells correlated with an increasing infiltration of CD68⁺ macrophages in liver metastatic lesions [80, 81], suggesting a role of macrophages in regulating the function of T cells in liver metastasis. In this part, we summarize the crosstalk between hepatic macrophages and T cells in liver metastasis. On one hand, macrophages contribute to an immunosuppressive environment by inhibiting the function of T cells through cell-to-cell interaction and the secretion of immunosuppressive factors [15, 82, 83] (Fig. 3). For example, in a CRC liver metastasis mouse model, GATA binding protein 6 (GATA6)⁺ peritoneal macrophages were recruited to the metastases site in the liver and subsequently up-regulated the expression of Programmed cell death 1 ligand 1 (PD-L1), a suppressive immune checkpoint, induced by apoptotic tumor cells, which resulted in inhibition of CD8⁺ T cell function [84]. Recently, several studies showed that the interaction between Secreted Phosphoprotein 1 (SPP1)⁺ macrophages and CD8⁺ T cells mediated by GDF15-TGFBR2 signaling or CD44 leads to exhaustion of CD8⁺ T cells and immunosuppression during liver metastasis, this effect can be reversed by neutralizing antibody of GDF15 and CD44 [82, 85]. Furthermore, liver metastases-infiltrated macrophages can induce activated T cells apoptosis via the Fas-Fas Ligand (FasL) pathway [86].Additionally, fatty acid-binding protein 7 (FABP7) was confirmed to overexpressing in macrophage, and correlating with poor prognosis of CRLM patients. Overexpressing FABP7 facilitates fatty acid uptake and accumulation in macrophages and further deliver lipids to CD8⁺ T cells resulting increase of PD-1 and TIM3 that are biomarker for T cell exhaustion [87]. On the other hand, macrophages help to create an immunosuppressive environment via inducing an inhibitory phenotype of T cells, such as Treg cells and Th17 cells. Th17 cells are predominantly enriched in CRC liver metastasis tissue and promoted liver metastasis by the TNF superfamily member 12 (TNFSF12 or TWEAK)- TNF receptor superfamily member 12 A (TNFRSF12A or Fn14) interaction. Importantly, CD163⁺ macrophages are responsible for the recruitment of Th17 cells and promote the differentiation of Th17 cells into Treg cells, leading to an immunosuppressive microenvironment in CRC liver metastasis [88]. In addition, tumor-resident microbiota enhances the production of lactate and mediates M2 macrophage polarization by suppressing nuclear factor-κB -gene binding (NF-κB) signaling through retinoic acid-inducible gene 1 (RIG-I) lactylation, which contributes to the inhibition of CD8⁺ T cells and increases the infiltration of Programmed cell death protein 1 (PD-1)⁺ Treg cells to facilitate CRC liver metastasis [89]. Moreover, Pharmacological inhibition of macrophages has been showed to inhibit liver metastasis by decreasing CD4⁺CD25⁺ Treg cells both in the blood and liver metastasis [90].

NK cells

NK cells are a critical subset of innate lymphoid cells that differentiate from the bone marrow progenitor cells. An essential function of NK cells is to recognize normal cells and cells in distress, including tumor cells, and to eliminate the latter directly or indirectly [91–93]. These immune effectors execute dual surveillance functions through: (I) direct cytotoxicity mediated by granzymes (GZMA, GZMB) and perforin secretion [92, 93], (II) immunomodulation via cytokine production including interferon-γ (IFN-γ), growth factors such as FMS-like tyrosine kinase 3 ligand (FLT-3 L), and granulocyte–macrophage colony-stimulating factor [93]. Importantly, emerging evidence suggests their unique capacity to suppress metastatic progression and control the development of tumor metastases by putting metastatic cells into a dormant state or killing CTCs prior to niche colonization [93, 94] (Fig. 3). The liver microenvironment harbors substantial NK cell populations under homeostatic conditions. While their precise role in hepatic metastasis remains incompletely characterized. Recent studies showed that macrophages play an important role in activating NK cells in liver metastasis. It has been reported that macrophage Stimulator of interferon genes protein (STING) signaling contribute to secretion of IL-18 and Interleukin-1β (IL-1β) by NOD-like receptor family pyrin domain containing 3 (NLRP3) activation in CRC live metastasis, which subsequently induce an antitumor phenotype of NK cells [95]. Furthermore, another study demonstrated the role of macrophages in activating NK cells during liver metastasis. This study revealed that increased infiltration of CD49a + NK cells in CRLM was significantly associated with a better prognosis. Further analysis indicated that F4/80^high macrophages mediated NK cell recruitment through CXCL9 secretion, which was validated using neutralizing antibodies against CSF1 and CCR2 to deplete macrophages. Additionally, TGF-β inhibitor interventions demonstrated that macrophages induced the CD49a + NK cell phenotype via TGF-β signaling [96]. Importantly, not only do macrophages have an effect on NK cells, but NK cells can also regulate the function of macrophages. In a previous study, Timmers M and colleagues have proved that KCs residing in liver sinusoids can capture tumor cells in circulation and phagocytose them, and this important function is dependent on NK cells because the depletion of NK cell leads to dramatic inhibition of the phagocytic ability of macrophages during liver metastasis [97].

Macrophages and hepatic parenchymal cells in liver metastasis

Hepatocyte

The liver is composed of parenchymal and non-parenchymal cell components. Hepatocytes, constituting over 70% of the hepatic parenchymal cell population, demonstrates remarkable functional complexity in liver metastasis regulation [98, 99]. Emerging evidence highlights hepatocytes as pivotal mediators of liver metastasis through multifaceted mechanisms. For instance, hepatocyte-derived plexin B2 interacts with class IV semaphorins expressed on disseminated tumor cells, driving epithelial-like transformation and facilitating metastatic colonization in the liver [100]. Furthermore, during metastatic progression, hepatocytes activate the IL-6-STAT3 signaling pathway to secrete serum amyloid A1/2 (SAA1/2), which establishes a pro-metastatic niche by recruiting macrophages and neutrophils to the tumor microenvironment [101]. Mechanistically, Wu and colleagues further revealed that hepatocyte-secreted SAA binds to formyl peptide receptor 1 (FPR1) on macrophages to promote their recruitment, while simultaneously interacting with Toll-like receptor 2 (TLR2) to induce M2 polarization of macrophages, thereby fostering an immunosuppressive microenvironment conducive to liver metastasis [102] (Fig. 3).

Macrophages and hepatic non-parenchymal cells in liver metastasis

Fibroblast/ HSCs

HSCs, residing in the perisinusoidal space of the liver, constitute essential components of hepatic non-parenchymal cells. These cells have been demonstrated to play pivotal roles in various liver pathologies including liver fibrosis [103], liver cancer [104], non-alcoholic steatohepatitis [105], and liver inflammation [106]. Recently, it was reported that HSCs activation-mediated liver collagen deposition facilitates liver metastasis by promoting the colonization of tumor cells [107]. However, in liver metastasis, HSCs activation may be attributed to macrophages, in which crosstalk between macrophages and HSCs has been described to promote liver metastasis. Importantly, the secretion of TGF-β by macrophages is the main manner to HSCs activation in liver metastasis. For example, KCs uptake pancreatic ductal adenocarcinoma (PDAC)-derived exosomes containing Macrophage migration inhibitory factor (MIF) to promote the expression and secretion of TGF-β in KCs and subsequently mediate HSCs activation to drive metastatic niche formation in the liver, which this effect can be blocked by TGF-beta type I receptor inhibitor (A83-01) [108]. Furthermore, in the splenic injection mouse model of liver metastasis, during the early phase defined as the first 7 days post-injection, gastric cancer-derived lipopolysaccharide binding protein (LBP) increased immune cell infiltration and induced HSCs activation to enhance collagen deposition in the metastatic microenvironment, thereby promoting liver metastasis formation. Further macrophage depletion using clodronate liposome demonstrated that macrophages mediate LBP-induced HSC activation. Mechanistically, LBP protein upregulated macrophage TGF-β secretion leading to activation of HSCs through TLR4-NF-κB signaling, and this effect was significantly reversed by the TGF-β receptor inhibitor Galunisertib [109]. Moreover, in PDAC liver metastasis, granulin-secreting macrophages are recruited to the liver and secrete granulin to activate HSCs, leading to collagen deposition in the liver metastatic niche [107]. In addition, the macrophage-fibroblast crosstalk also induces an immunosuppressive microenvironment in liver metastasis. In pancreatic cancer, Progranulin (PGRN)-mediated communication between macrophages and fibroblasts can enhance the expression of SPP1 encoding osteopontin protein, to induce immunosuppression in liver metastasis [110]. During the process of steatotic liver-promoted CRLM, the fibrotic microenvironment is characterized by increased cancer-associated fibroblast numbers and collagen deposition, along with significant accumulation of hyaluronic acid (HA). Mechanistic studies further revealed that HA synthase 2 (HS2A) upregulation drives HA production and secretion, which induces immunosuppression of macrophages and T cell dysfunction through a CD44 membrane receptor-dependent manner, manifested by elevated expression of PD-L1 and TIM3. Importantly, these effects were reversed by conditional knockout of HS2A in HSCs and inhibition of HA synthesis with inhibitor 4-methylumbelliferone (4-MU) [111] (Fig. 3).

Therapeutic strategies targeting macrophage for liver metastasis intervention

Surgical resection remains the primary treatment for liver metastasis to prolong patient survival; however, only a small minority of patients can benefit from surgical resection, significantly limiting therapeutic options and overall efficacy. For patients who are ineligible for resection, systemic therapy serves as a critical approach to extend survival and improve quality of life. Nevertheless, treatment strategies vary depending on the tumor type underlying the liver metastasis. For example, 5-fluorouracil combined with irinotecan or oxaliplatin constitutes the primary regimen for CRLM, whereas for non-CRLM, tyrosine kinase inhibitors (TKIs) such as sunitinib and imatinib represent key therapeutic options. A major challenge associated with TKI therapy, however, is the development of drug resistance, as many patients eventually develop resistance to agents such as imatinib upon disease progression. Given the pivotal role of macrophages in liver metastasis progression, emerging evidence suggests that therapeutic targeting of hepatic macrophages may constitute a viable strategy for liver metastasis intervention. According to current studies, hepatic macrophage-targeted therapeutic strategies for liver metastasis can be mainly classified into four approaches: (I) Pharmacological depletion of KCs and Mo-Macs by using agents such as clodronate-encapsulated liposomes [45, 57, 59, 77, 90, 112]; (II) Genetic ablation of macrophage-specific targets via conditional knockout models [49, 51, 58, 85].; (III) Pathway-selective inhibition using small-molecule antagonists [15, 48, 56, 80, 82, 87–89, 113]; and (IV) Neutralizing antibodies against secreted signaling molecules or membrane receptors [16, 46, 55, 73, 84, 95] (Table 2).

Table 2.

Targeting macrophage for liver metastasis therapeutic

Type of intervention	Interventions	Cancer Type	The role/phenotype of macrophages	Target	Cell interaction	Combination therapy	Reference
Depletion of macrophage	Clodronate liposome	CRC	Recruitment and M2 polarization of Mac	Macrophage	Tumor cells-Mac	—	[57]
		CRC	CXCL16 mediate anti-tumor phenotype of macrophages		Mac-Tumor cells	—	[59]
		CRC	Recruitment and M2 polarization of Mac		Mac-Tumor cells	—	[45]
		CRC	Mediating T cell apoptosis		Mac-T cells	α-PD-L1	[86]
		CRC	Mediating secretion of TGFβ		Tumor cells-Mac	—	[112]
		Multiple tumor	Phagocytize tumor cells		KCs-Tumor cells	—	[77]
		PDAC	Promoting CD4 + CD25 + T cells infiltration		Mac-T cells	—	[90]
	GdCl₃	CRC	Inhibiting liver metastasis		—	—	[76]
	Anti-CSF1R	CRC	Secreting CXCL9 to recruit NK cells		Mac-NK cells	—	[96]
		Multiple tumor	Engulfing tumor cells		KCs-Tumor cells	—	[17]
Neutralizing antibody	Anti-FcγRIV	Melanoma	Phagocytosis of tumor cells	FcγRIV	KCs-Tumor cells	—	[73]
	Anti-4-1BBL	CRC	Promoting antitumor activity of NK cells	4-1BBL/4-1BB	Mac-NK cells	—	[95]
	Anti-CXCL13	CRC	Recruitment and M2 polarization of Mac	CXCL13-CXCR5	Tumor cells-Mac	—	[46]
	Anti-PD-L1	CRC、breast cancer、Melanoma	Inhibiting T cells function	PD-L1-PD-1	Mac-T cells	—	[84]
	Anti-CTHRC1	CRC	Recruitment and M2 polarization of Mac	CTHRC1/TGF-βR	Tumor cells-Mac	α-PD-1	[55]
	Antibody(Jagged1、IL-1a、CCL2、uPAR、IL-6)	PADC	Recruitment and M2 polarization of Mac	Jagged1、IL-1a、CCL2、uPAR、IL-6	Tumor cells-Mac	—	[16]
Knockou of genes	siRNA(MMP9)	CRC	Increasing vascular permeability	MMP9	Mac-vascular	—	[65]
	Ca5R-KO	CRC	Recruitment and M2 polarization of Mac	Ca5R	Tumor cells-Mac	—	[51]
	CD36-CKO(Mac)	Lung cancer、Melanoma、CRC	Recruitment and M2 polarization of Mac; Inhibiting T cell function	CD36	Tumor cells-Mac; Mac-T cells	—	[49]
	YAP-KO	CRC	Recruitment and M2 polarization of Mac	YAP	Tumor cells-Mac	—	[124]
	Ndrg2-KO	CRC	M2 polarization of Mac	Ndrg22	Tumor cells-Mac	—	[118]
	CD93-CKO	CRC、PDAC	M2 polarization of Mac	CD93	Tumor cells-Mac	α-PD-1	[58]
	SPP1-KO	Melanoma、CRC	Inhibiting T cell function	SPP1	Mac-T cells	—	[85]
Inhibitors	HDAC (TMP195)	CRC	Increasing vascularization	HDAC	Mac-vascular	—	[122]
	MerTK (UNC2250)	PDAC	Mediating immunosuppression	MerTK	Mac-T cells	—	[80]
	GDF15(HY-P99241)	Gastric cancer	Inhibiting T cell function	GDF15	Mac-T cells	—	[82]
	CCR5 inhibitor (Maraviroc)	CRC	Recruiting Th17 cells	CCR5	Mac-T cells	—	[88]
	RIG-I lactylation‌ (AJ-64234603004)	CRC	Inhibiting T cell function; promoting PD-1 expression of Treg cells	RIG-I	Mac-T cells	5-fluorocrail (5-FU)	[89]
	VEGFR (KRN-633)	Gastric cancer	Recruitment and M2 polarization of Mac	VEGFR	Tumor cells-Mac	—	[48]
	Syk(piceatannol, BAY61-3606 )	CRC	M2 polarization of Mac	Syk	Tumor cells-Mac	—	[56]
	CCR2 (PF-04136309)	CRC	Inhibiting T cell function	CCR2	Mac-T cells	—	[15, 83]
	SPHK1 (PF543)	CRC	Promoting tumor mediated TAM recruitment	SPHK1	Mac-Tumor cells	α-PD-1	[113]
	FABP7 (SBFI-26)	CRC	Inhibiting T cell function	FABP7	Mac-T cells	—	[87]
Other	Targeting through bacteria	Melanoma, CRC	Inhibiting liver metastasis	KCs	KCs-Tumor cells	—	[70]
	Gene transduction	Melanoma	Inhibiting liver metastasis	Macrophage	Mac-Tumor cells	—	[120]
	β-glucan	PDAC	Activating T cells	KCs	KCs-T cells	α-PD-1	[74]
	Bufalin	CRC	M2 polarization of Mac	Macrophage	Tumor cells-Mac	—	[54]

However, these intervention strategies still have some limitations and challenges. Firstly, accumulating evidence from multiple studies and single-cell sequencing analyses reveals significant heterogeneity among hepatic macrophages, particularly in liver metastatic microenvironments [114–117]. Different macrophage subpopulations may exhibit opposing functional roles in either promoting or suppressing liver metastasis. For instance, Donadon and colleagues showed that macrophages exhibit distinct morphological characteristics in CRLM. Compared to small tumor associated macrophages (S-TAMs), large tumor associated macrophages (L-TAMs) display significantly larger cellular area and diameter, accompanied by enrichment of lipid metabolism-related signaling pathways, and are associated with 5-year disease-free survival rates of 27.8% and 0.2%, respectively [115]. Although clodronate-based depletion effectively eliminates macrophages in the liver metastatic environment, conflicting experimental outcomes have often been reported across studies, since this approach non-selectively removes both anti-metastatic and pro-metastatic macrophage subsets. Secondly, targeted gene knockout is widely recognized as a pivotal therapeutic intervention strategy. For example, C5aR, a pivotal receptor for tumor-derived C5a, orchestrates macrophage recruitment during hepatic metastasis. And, transplantation of wild-type bone marrow-derived macrophages (BMDMs) into C5aR knockout (C5aR-KO) mice resulted in dramatically accelerated liver metastasis progression, whereas C5aR-KO effectively attenuated metastasis through impaired leukocyte trafficking [51]. Furthermore, CD36 mediates tumor-derived lipid-induced M2 polarization of hepatic macrophages. Conditional CD36 knockout in macrophages not only prevented metastasis-associated M2 polarization but also activated anti-tumor immune responses [49, 50]. Additionally, N-myc Downstream Regulated Gene 2 (Ndrg2)-KO can inhibit the progression of liver metastasis through activating M1-polarization of macrophage [118]. In spite of conditional knockout of macrophage-specific targets demonstrates enhanced intervention precision, current applications remain largely confined to experimental mouse models, thereby limiting clinical translation. Therefore, it is worthwhile and challengeable to develop more practical and efficient delivery systems of gene intervention drugs. Thirdly, small-molecule inhibitors and neutralizing antibodies targeting to specific targets have shown promising anti-metastatic efficacy in mouse models by modulating macrophage phenotypes or blocking intercellular signaling. Emerging pharmacological strategies demonstrate therapeutic potential through hepatic macrophage modulation. Small-molecule inhibitors targeting key regulatory targets including Tyrosine-protein kinase Mer MerTK (UNC2250), GDF15 (HY-P99241), CCR5 (Maraviroc), HDAC (TMP195), CSF1R (PLX5622), SyK (Piceatannol), and VEGF (KRN-633) have shown efficacy in suppressing liver metastasis by specifically inhibiting their respective macrophage-expressed targets. Furthermore, neutralizing antibodies against macrophage-derived factors (CXCL13 [46], CTHRC1 [55], IL-1α, CCL2, uPAR, IL-6 [16], tumor necrosis factor ligand superfamily (TNFSF9 or 4-1BBL) [95], FcγR IV [73]) effectively attenuate metastatic progression via paracrine signaling blockade. Notably, PD-L1 checkpoint inhibition using anti-PD-L1 monoclonal antibodies restores anti-tumor immunity by reversing T-cell exhaustion induced by PD-L1high hepatic macrophages [84], significantly reducing metastatic burden. However, these drugs lack cellular specificity, which affect all cells expressing the targeted molecules and may induce unpredictable off-target effects.

Furthermore, because of the inherent phagocytic capacity of macrophages, exogenously administered therapeutic agents can be preferentially taken up by hepatic macrophages [26]. This characteristic can be leveraged to develop macrophage-targeted therapeutic strategies for the treatment of liver metastasis. Zhou and colleagues utilized the phagocytic capacity of macrophages and encapsulated siRNA of the phagocytic receptor MerTK (siMerTK) into a clinically approved lipid nanoparticle platform (LNP) to treat liver metastasis in mouse model. In their study, the LNP-siMerTK were mainly up-taken by hepatic macrophages and polarize macrophages into anti-tumor phenotype by blocking efferocytosis [119]. Moreover, Liu et al. take advantage of the superior capacity of KCs to capture circulating bacteria and developed a delivery system that administration of Escherichia coli producing clustered regularly interspersed short palindromic repeats CRISPR machinery enables efficient editing of genes of interest in KCs and this technique acquired exciting therapeutic effect in liver metastasis treatment [70]. Thomas et al. found that β-glucan as a pathogen-associated molecular pattern (PAMP) can be up-taken by macrophages to enhance the anti-tumor effect in liver metastasis through increasing phagocytosis of tumor cells [74].In addition to interventional strategies utilizing the phagocytic properties of macrophages for targeted delivery, engineered modifications to restore their anti-tumor activity also represent a novel approach in macrophage-targeted therapies for liver metastasis. A new study demonstrates that arming liver and tumor-associated macrophages in vivo to co-express tumor antigens (TAs), IFNα, and IL-12 unleashes robust anti-tumor immune responses, leading to the regression of liver metastases [120]. In a word, macrophages may be a prospective target for liver metastasis intervention.

Conclusion and perspective

Liver metastasis is driven by multiple biological processes, including pre-metastatic niche formation, tumor cell colonization and outgrowth of tumor cell in the liver, which is largely regulated by the liver metastatic microenvironment. The liver is generally regarded as an immunotolerant organ, harboring a unique immunosuppressive microenvironment, that facilitates the dissemination and colonization of metastatic tumors [25]. This intrinsic property is due to its continuous exposure to diverse antigens, such as gut-derived bacterial components and xenobiotics, which enabling hepatic immune cells to phagocytize and clear pathogens while avoiding disproportionate immune activation. This immunological landscape of the liver is further remodeled by tumor-derived factors to establish appropriate soil for DTC. Prior to liver metastasis formation, tumor-derived cytokines, exosomes, and other mediators remodel the liver microenvironment to establish a pre-metastatic niche [15, 16, 45–51]. Following DTC colonization, interactions with liver-resident cellular components including LSECs, KCs and HSCs, to drive tumor cell colonization and outgrowth, extracellular matrix remodeling, and subsequent metastatic progression [121, 122]. However, the inherent cellular diversity and plasticity within the hepatic microenvironment collectively make it difficult for intervening in liver metastasis [114–116]. Furthermore, the optimal therapeutic window proves particularly challenging to define due to both the multistage progression of liver metastasis and the variable therapeutic efficacy across different phases. Therefore, identifying core responsive cells during liver metastasis progression and elucidating their pivotal regulatory roles will provide practical insights for precise intervention strategies of liver metastasis therapy.

Macrophages, as key cellular components of the liver microenvironment, play a pivotal role in modulating the liver tumor microenvironment. This review comprehensively analyzes the multifaceted roles of hepatic macrophages in liver metastasis. During the early metastatic phase, tumor-derived factors recruit Mo-Macs into the liver and regulate the polarization phenotypes of both Mo-Macs and KCs, thereby mediating the formation of premetastatic niche [121]. Additionally, macrophages directly interact with tumor cells to promote their colonization and survival. Furthermore, in this review, we discuss how macrophages crosstalk with other immune cells (e.g., T cells, NK cells) [80, 82, 95] and non-immune cells (e.g., hepatic stellate cells, hepatocytes) [102, 110], leading to T-cell dysfunction and collagen deposition-mediated extracellular matrix remodeling, which collectively reprogram the tumor microenvironment. These findings collectively suggest that macrophages may serve as central regulators in liver metastasis progression, and targeting macrophages could represent a promising therapeutic strategy. However, recent studies employing single-cell sequencing and flow cytometry have revealed substantial heterogeneity among hepatic macrophages, which can be categorized into distinct functional subsets [114–116]. This heterogeneity poses challenges for macrophage-targeted therapies, as precise interventions selectively inhibiting pro-tumorigenic subsets while preserving anti-tumor populations are required. Moreover, the growing list of identified macrophage-associated targets, such as PD-L1, CTHRC1, CCL2 and CCR2, necessitates further clinical validation to determine which therapeutic targets may confer clinical benefits in patients [15, 55, 83, 84]. However, a search using “liver metastasis” as a keyword on web (https://clinicaltrials.gov) revealed that there are still very few clinical trials investigating drug-targeted therapies for liver metastasis, indicating that the clinical translation of targeted theraputics for liver metastasis continues to face significant challenges. In-depth investigation to elucidate the central regulatory role of macrophages in liver metastasis will facilitate the development of novel strategies targeting macrophages for liver metastasis therapy (Tables 2 and 3).

Table 3.

Completed clinical trials associated with liver metastasis

NCT Number	Study Title	Cancer Type	Interventions	Phases	Study Type
NCT04338685	A Study Evaluating Safety, Pharmacokinetics, Pharmacodynamics, And Clinical Activity of RO7119929 (TLR7 Agonist) in Participants with Unresectable Advanced or Metastatic Hepatocellular Carcinoma, Biliary Tract Cancer, or Solid Tumors with Hepatic Metastases	CRC	Drug: RO7119929, Drug: Tocilizumab	Phase1	Interventional
NCT04463368	Isolated Hepatic Perfusion in Combination with Ipilimumab and Nivolumab in Patients with Uveal Melanoma Metastases	CRC	Procedure: Isolated hepatic perfusion, Drug: Ipilimumab, Drug: Nivolumab	Phase1	Interventional
NCT05183776	Clinical Validation of a Fractional Administration Device for Holmium-166 SIRT	CRC	Device: Fractional administration Device	NA	Interventional
NCT05616039	I-FIGS Feasibility Study	CRC	Procedure: Standard Liver Surgery, Diagnostic Test: Indocyanine Fluorescent Image Guided Surgery (I-FIGS)	NA	Interventional
NCT04676633	Safety, Tolerability, PK, Anti-Tumor Activity of STP705 in Subjects with Advanced/Metastatic or Surgically Unresectable Solid Tumors Who Are Refractory to Standard Therapy	CRC	Drug: STP705	Phase1	Interventional
NCT04942665	Low Dose ICG for Biliary Tract and Tumor Imaging	CRC	Drug: Indocyanine green, Device: PINPOINT Endoscopic Fluorescence	Phase2	Interventional
NCT05995990	Raman Spectroscopy for Liver Tumours Following Liver Surgery	CRC	Device: Raman Spectrometry	NA	Interventional
NCT04880772	Clinical Trial Comparing Standard Care Versus Prehabilitation in Patients Undergoing Cancer Surgery	CRC	Other: Prehabilitation	NA	Interventional
NCT06184841	HAIC Combined Sintilimab for Liver Metastasis from Esophageal Squamous Cell Carcinoma	CRC	Drug: HAIC combined with intravenous PD-1	Phase1	Interventional
NCT05169957	Hepatic Ablation of Melanoma Metastases to Enhance Immunotherapy Response, a Phase I Clinical Trial (HAMMER I)	CRC	Drug: Ipilimumab, Drug: Nivolumab, Procedure: Stereotactic Body Radiation Therapy	Phase1	Interventional
NCT04517643	TheraSphere for Treatment of Metastases in Liver	CRC	Other: Therasphere Therapy	NA	Interventional
NCT05293041	Argipressin’s Influence on Blood Loss During Hepatic Resection	CRC	Drug: Argipressin, Drug: Placebo	Phase4	Interventional
NCT03993327	An Imaging Agent (I-124 M5A) in Detecting CEA-Positive Liver Metastases in Patients with Colorectal Cancer	CRC	Biological: Iodine I 124 Monoclonal Antibody M5A, Procedure: Positron Emission Tomography	Phase1	Interventional
NCT04150731	16α-18 F-fluor-17β-estradiol PET/CT for Visualisation of Estrogen Receptor Positive Liver Metastases from Breast Cancer	Breast Cancer	Radiation: 16α-18 F-fluor-17β-estradiol	Phase1, Phase2	Interventional
NCT05182112	Radiation Therapy (RT) and Chemo-therapy for the Treatment of Pancreatic Cancer with Homologous Recombination Deficiency That Has Spread to the Liver	PDAC	Radiation: Whole liver irRadiation (WLI), Drug: Gemcitabine and Cisplatin	Phase1	Interventional

In this review, we have summarized that hepatic macrophages can interact with T cells, NK cells, hepatocytes, and HSCs to promote liver metastasis. In the future, the crosstalk between hepatic macrophages, other immune cells, and hepatic stromal cells should be clarified. For instance, neutrophil infiltration is increased in the liver during liver metastasis, and whether macrophage-neutrophil crosstalk is important for liver metastasis remains unclear [52, 123]. In summary, hepatic macrophages are a vital component of liver metastasis, and further research is needed to elucidate the exact and precise effects on liver metastasis.

Author contributions

Conceptualization, F.Z., S-P. Y., W-H W.; original draft preparation, W-H.W. and Z-Y.Y.; editing and review, H-D. C., Y-D. H, L-H. J., S-P.Y., Y. L and F.Z.; funding acquisition, W-H.W., Z-Y. Y and S-P. Y. All authors have read and agreed to the published version of the manuscript.

Funding

This study was supported by National Natural Science Foundation of China (No.82403610), New Chongqing Youth Innovation Talent Program (No. CSTB2024NSCQ-QCXMX0066), Medical Research Project of Chongqing Municipal Health Commission (No. 2025WSJK089) and Natural Science Foundation of Chongqing Municipality (No. cstc2021jcyj-msxmX0622).

Data availability

Not applicable.

Declarations

Ethical approval

Not applicable.

Consent to participate

Not applicable.

Conflict of interest

The authors declare no conflict of interest.