Cancer stem cell-immune cell crosstalk in the tumor microenvironment for liver cancer progression

Abstract

Crosstalk between cancer cells and the immune microenvironment is determinant for liver cancer progression. A tumor subpopulation called liver cancer stem cells (CSCs) significantly accounts for the initiation, metastasis, therapeutic resistance, and recurrence of liver cancer. Emerging evidence demonstrates that the interaction between liver CSCs and immune cells plays a crucial role in shaping an immunosuppressive microenvironment and determining immunotherapy responses. This review sheds light on the bidirectional crosstalk between liver CSCs and immune cells for liver cancer progression, as well as the underlying molecular mechanisms after presenting an overview of liver CSCs characteristic and their microenvironment. Finally, we discuss the potential application of liver CSCs-targeted immunotherapy for liver cancer treatment.

Keywords

liver cancer / cancer stem cell / immune cell / immunotherapy

1 Introduction

Liver cancer, primarily hepatocellular carcinoma (HCC), is the sixth most common aggressive cancer [1]. Currently, several kinds of molecular targeted drugs have been approved for HCC treatment, including kinase inhibitors, angiogenesis inhibitors and immune checkpoint inhibitors, which only provide modest benefits [2]. Previous studies indicated that the main manifestations of malignant tumors related to the existence of cancer stem cells (CSCs) [3]. Emerging evidences show that a subset of stem cells or precursor cells in HCC, namely liver CSCs [4–6]. Heterogeneous HCC is largely explained by liver CSCs expressing different surface markers. Alterations in the tumor microenvironment (TME) are also known to promote tumor cell heterogeneity [7]. The liver CSC TME is essential for maintaining the properties of CSCs, protecting them from being attacked by the immune system and maintaining their metastatic potential [8]. Another confounding variable is the interaction of the immune cell lineage with CSCs, as both immune evasion and CSCs are considered integral parts of tumor growth and metastatic spread [9]. Harnessing the immune system to recognize and efficiently eliminate tumor cells has been successfully used in HCC treatment.

This review summarizes current knowledge of liver CSCs, and highlights the impact of the microenvironment niche on HCC stemness. Importantly, we focus on the significance of crosstalk between CSCs and the immune system, and discuss the potential application of liver CSCs-targeted immunotherapy in liver cancer therapy.

2 Liver CSCs characteristic

2.1 The origin of liver CSCs

The origins of liver CSCs are controversial. One potential origin of CSCs is from liver stem/progenitor cells, which is concluded from the fact that liver CSCs share a massive number of similar characteristics with normal stem cells [10-13]. Hepatic progenitors expressing H-Ras and SV40LT proteins tend to transform into liver CSCs [14]. A leading cause of tumor cells acquiring CSC-like features is also thought to be the reprogramming and dedifferentiation of non-CSCs [14,15]. Another view is that liver CSCs may derive from the dedifferentiation of mature hepatocytes [16–18]. Chronic liver disease causes recurrent inflammatory response and regeneration of hepatocytes, resulting in genetic and epigenetic alterations in hepatocytes and hepatic progenitor cells, which also contribute to liver CSCs formation [14,19]. Additionally, the origin of liver CSCs is also investigated by lineage-tracing studies and ablation of CSC markers (namely, epithelial cell adhesion molecule (EpCAM) and LGR5) in intact mouse HCC tumors. EpCAM⁺ ductal cells are the main cell source of liver cancer due to inflammation-associated tumorigenesis in a mouse model of HCC [20]. Furthermore, LGR5⁺ hepatocytes are found to be responsible for diethylnitrosamine (DEN)-induced HCC and highly sensitive to neoplastic transformation triggered by ERBB pathway activation [21]. Collectively, these studies suggest that liver CSCs originate from stem/progenitor cells, proliferating ductal cells and hepatocytes.

2.2 Cell surface markers of liver CSCs

Although the pathogenesis of HCC remains unclear, liver CSCs are vital for HCC formation and exhibit heterogeneity, which due to the presence of tumor cell subsets expressing different markers in HCC [22–24]. Several surface markers for liver CSC subpopulations (CD133 [25], CD90 [26], CD44 [27], EpCAM [28], CD13 [29], oval cell marker (OV-6), CD47 [30], delta like 1 homolog (DLK1) [31], CD24, intercellular adhesion molecule 1 (ICAM1) [32], α2δ1 [33], and keratin 19 (CK19) [34]) in HCC have been identified. Recent studies indicate that CD133 regulates neurotensin, interleukin (IL)-8, CXCL1, and MAPK signaling to maintain the characteristics of CSCs [25,35,36]. Additionally, CD90⁺ and CD44⁺ HCC cells also display tumorigenic and metastatic potential [26,37]. Recently, CD44 has been found to regulate TGF-β-induced epithelial-mesenchymal transition (EMT) [27] and maintain CSCs properties via regulating redox status [38]. In addition, EpCAM⁺ HCC cells show CSC traits that are highly invasive and tumorigenic [28,39]. Moreover, CD13 regulates the reactive oxygen species (ROS) scavenger pathway to prevent cells from apoptosis [29]. CD47 modulates the cathepsin S/protease-activated receptor 2 paracrine pathway to regulate HCC stemness [30]. DLK1⁺ cells possess robust chemoresistance and self-renewal potential in vitro and tumor-initiating capacity in vivo [31]. Both CD24 and ICAM1 are functional liver CSC markers that activate stemness-associated factor Nanog to initiate HCC [32]. Furthermore, α2δ1 has been demonstrated to potentiate liver CSCs characteristics by activating pro-survival pathways in a calcium-dependent mechanism [33]. CK19 is also a functional CSC marker that contributes to tumor progression [34,40]. Overall, these results propose that liver CSC markers provide a promising therapeutic target for overcoming HCC treatment resistance.

2.3 Liver CSCs-associated signaling pathway

Liver CSCs mediate therapeutic resistance and relate with the poor prognosis of HCC patients [41,42]. Intricate liver CSC-associated signaling pathways are generally related to HCC development [43]. Evidence is emerging to support that Wnt/β-catenin signaling is activated in different liver CSC subsets [44–47], including EpCAM⁺ [48], CD133⁺ [49], OV6⁺ [50], Sox9⁺ [51], and Lgr5⁺ [30] liver CSCs. Shp2 [52] and EPHB2 [53] favor liver CSC extension through β-catenin signaling. Of note, EpCAM is identified as a direct target of β-catenin, and Wnt/β-catenin signaling activates the liver CSC marker EpCAM [54]. AKT and AKT/GSK3β signaling is often altered in liver CSCs, which are related to the Wnt/β-catenin signaling. Accordingly, EPHB2 regulates cancer stemness and drug resistance through AKT/GSK3β/β-catenin axis [55]. The NOTCH signaling is hyperactive and plays physiologic roles in HCC. It is also involved in stem cell self-renewal and differentiation [56,57]. It is reported that C8orf4 decreases the proportion of CD13⁺CD133⁺ liver CSCs by inhibiting NOTCH2 signaling [58]. Consistently, CD133⁺ liver CSCs have a higher NOTCH expression [49,56]. Moreover, iNOS/nitric oxide (NO) activates NOTCH1 via regulating TACE/ADAM17 signaling in liver CSCs to promote hepatocarcinogenesis [59]. IL-6/STAT3 activation in liver CSCs is also positively correlated with HCC development [60]. Hepatitis B virus X (HBx) activates IL-6/STAT3 signaling to facilitate the carcinogenicity of hepatic progenitor cells [61]. Additionally, STAT3 upregulates Nanog expression to promote the extension of CD24⁺ liver CSCs [62]. Long-term tobacco smoke exposure enhances HCC cell stemness properties through IL-33/p38 axis [63]. Furthermore, the JAK/STAT pathway inhibits stem cell homeostasis by suppressing SP/CD44⁺ CSCs properties in HCC [64]. Consequently, distinct signaling pathways are involved in regulating liver CSC properties in HCC, which provides various insights for studying the pathogenesis of HCC.

3 Tumor microenvironment of liver CSCs

HCC is a highly heterogeneous disease, which forms an immune tolerance microenvironment. CSCs exist in the TME, maintaining an undifferentiated state and self-renewal potential [7]. Emerging evidence indicates the potential role and influence of the liver CSC microenvironment on modulating HCC tumorigenesis, EMT and tumor progression [65]. The liver CSC microenvironment encompasses a series of tumor-related components, such as tumor parenchymal cells (containing non-CSC cancer cells and CSCs), stromal cells [66], immune cells, cytokines, extracellular vesicles (EVs) [67], extracellular matrix (ECM), oxygen, nutrients [68], and hepatitis virus infection [69,70], which in turn promote tumor progression (Fig.1).

In addition to the tumor parenchymal cells, the stromal cells and their secreted cytokines are the main components that constitute the TME. It is acknowledged that cancer-associated fibroblasts (CAFs) regulate the stem cell properties and tumorigenicity of CSCs in several malignant tumors [71]. CAFs drive NOTCH3 expression to regulate the self-renewal of liver CSCs through LSD1 activation [72]. Subsequently, CAFs are able to trigger HCC stem cell-like characteristics via IL-6-STAT3-NOTCH signaling [73]. Our previous study found that tumor-derived exosomal miR-1247-3p stimulates the CAF to product IL-6 and IL-8 and exert the cancer cells to an EMT phenotype [66]. The findings suggest that CAFs interact with HCC to maintain and progress liver disease. Additionally, endothelial cells are essential components of the TME and contribute to tumor metastasis and angiogenesis [74]. CD133⁺ liver CSCs secrete IL-8 to promote angiogenesis through its effects on endothelial cells in the TME [35]. Moreover, CD90⁺ liver CSCs modulate the phenotype of endothelial cells by releasing exosomes containing H19 long non-coding (lncRNA) [75], implying the interaction between angiogenesis and liver CSCs in TME contributes to HCC development [76]. Taken together, these results suggest the critical roles of the stromal cells in regulating liver CSCs properties.

Immune cells are essential in driving CSC extension and CSC-specific avoidance of immunosurveillance. Molecules secreted by immune cells regulate cancer cell properties in the TME [77]. These inflammatory factors generate a CSC-inducing niche. It has been found that tumor-associated macrophages (TAMs) induce EMT and CSC-like properties through IL-6, TNF-α, and TGF-β1 in HCC [78–80]. Conversely, CSCs could also regulate immune cell antitumorigenic properties. CD47⁺ liver CSCs can escape phagocytosis by M1 macrophages in the TME [81,82]. Moreover, chemokine ligand 20 (CCL20) derived from hypoxia-induced liver CSCs lead to the recruitment of IDO⁺ TAMs and the inhibitory effect of T cell responses [83]. In addition, tumor-associated neutrophils (TANs) are essential in driving liver CSC expansion by secreting bone morphogenetic protein 2 (BMP2) and TGF-β2 [84]. TAN-induced HCC stem cell like cells generate more CXCL5 to recruit more TANs infiltration, which forms a positive feedback loop [84]. Moreover, immune suppressive cytokines secreted by myeloid derived suppressor cells (MDSCs) and regulatory T cells (Tregs) mainly mediate the formation of an immunotolerant niche [85]. Various tumors including HCC is infiltrated by Tregs [86]. Tregs inhibit the activities of effector T cells and avoid self-tolerance by secreting cytokines or direct contact. It has been investigated that Tregs are capable of promoting the invasion of HCC cells through TGF-β1-mediated EMT [87]. Liver CSCs can promote VEGF production, and thus inducing the recruitment of MDSCs in the TME [88]. Additionally, dendritic cells (DCs) could increase the apoptosis ratio of liver CSCs and exert the T cell-mediated antitumor immune response [89,90]. The ablation of Arf1 in CSCs could activate DCs and enhance T cell infiltration to potentiate anti-tumor immune response [90]. Accumulating evidence has demonstrated that liver CSCs contribute to the evasion of immunosurveillance by altering the phenotype of DCs and impairing their recruitment [91,92]. These investigations indicate the critical roles of immune cells in favoring the CSC niche.

T cells and natural killer (NK) cells are vital components of immune detection and play an essential role in long-term antigen-specific immune recognition [9]. Notably, they are able to directly identify and kill cancer cells. Interactions between CSCs and CD8⁺ T cells could be divided into two aspects: CSCs evade T cell-mediated cytotoxicity and inhibit the anti-tumor properties of T cells [9]. Programmed cell death 1 ligand 1 (PD-L1) expression on HCC cells interact with programmed cell death 1 (PD-1) on T cell to escape T cell-mediated immune surveillance. Interferon γ (IFN-γ) is a key factor secreted by the immune system to eliminate cancer cells [93]. IFN-γ derived from T cell could upregulate PD-L1 expression on HCC cells [94]. CSCs could impair cytotoxic T cell activity by altering their PD-L1 expression [95]. For example, the expression of PD-L1 is elevated in CD44⁺CD24⁺ALDH⁺ CSCs [96]. Moreover, CSCs decrease the expression of MHC-I to limit the presentation of neoantigens to T cells [97]. Additionally, liver CSC-derived cytokines can impair NK cell activity, such as circular ubiquitin-like with PHD and ring finger domain 1 RNA (circUHRF1) and granulin-epithelin precursor (GEP) [98,99].

ECM remodeling is critical for tumor progression and metastasis. Notably, ECM stiffness is vital for regulating liver CSC differentiation [100]. Matrix stiffness is found to promote a higher proportion of CD133⁺ or EpCAM⁺ HCC cells through integrin β1-activated mTOR signaling [101]. It is known that HCC cells remodel the ECM by upregulating ECM components and matrix metalloproteinases (MMPs). Nevertheless, CD133 expression in liver CSCs induce the production of MMP2 and metalloproteinase 9 to degrade ECM and promote immune escape by reducing their sensitivity to NK cells [102]. Moreover, liver CSCs produce lysyl oxidase (LOX) to promote the formation of a stiff ECM, thus enhancing the stiffness-mediated cancer stemness [103].

Hypoxia and glucose deprivation are also the main factors that stimulate vascular proliferation and promote HCC development [104]. HCC is a solid tumor and the tumor body grows rapidly, which forms a hypoxic environment. Hypoxia promote HCC cells proliferation [105], cause angiogenesis, and accelerate invasion [106]. For instance, HIF1α upregulates USP22 and TP53 at the transcriptional level under hypoxic conditions, and then promotes the production of CD24⁺ or CD44⁺ liver CSCs in HCC [107]. Similarly, the positive feedback loop between HIF-1α and SENP1 contributes to CD24⁺ subpopulation in HCC under hypoxia conditions [104]. Accordingly, it is necessary to clarify the changes in hypoxia-related response molecules in HCC cells. In addition, glucose deprivation generally appeared in large-growing tumors microenvironment such as HCC. Nutrient deprivation impacts the expansibility of liver CSCs [108]. Under low glucose conditions, hexosamine biosynthetic pathway (HBP) increases CD133⁺ subpopulation and enhances CD133 expression in HCC cells, leading to a more aggressive HCC progression [109]. Moreover, FUT1-mediated fucosylation drives HCC stemness through CD147, ICAM-1, EGFR, and EPHA2 via AKT/mTOR/4EBP1 axis [68]. In summary, hypoxia and glucose deprivation significantly affects liver CSCs maintenance.

Hepatitis B virus (HBV) [110] and hepatitis C virus (HCV) infection [111] are the largest risk factors for HCC occurrence and development, indicating that hepatitis virus-induced chronic liver inflammatory environment promotes hepatocarcinogenesis [112,113]. HBV PreS1 upregulates CSCs-related markers (CD133 and CD90) expression and facilitates the capacity of CSCs in HCC [69]. Additionally, HBx upregulates miR-5188 expression to promote β-catenin nuclear translocation and HCC stemness [114]. Similarly, HCV and HCFD mediate TLR4-NANOG and the leptin receptor (OBR)-STAT3 activation to accelerate HCC progression [115]. Moreover, Sal-like protein4 (SALL4) gene re-expression is induced by DNA demethylation in HCC cells, which is associated with HBV or HCV infection [116]. In brief, hepatitis virus infection contributes to the self-renewal of liver CSCs and facilitates HCC progression.

4 Liver CSCs-immune cell crosstalk in tumor progression

Immune cells are prominent components in modulating the CSC subpopulation. Previous studies have shown the relationship between liver CSCs and immune cells. CSCs can promote tumor initiation and progression, revealing that they have the advantage of evading immune surveillance. Emerging evidence support that some primary immune cells are necessary for CSCs expansion and enable them to escape immune surveillance via distinct mechanisms [9].

4.1 Protumorigenic immune cells

Protumorigenic immune cells mainly refer to tumor-associated myeloid cells in the innate immune system, including macrophages, granulocytes, and monocytes [117,118]. Extensive studies support that myeloid cells interact with HCC cells and affect the TME to regulate cancer progression and recurrence [119]. Therefore, the interplay between myeloid cells and adaptive immunity is also an essential modulator of tumor progression, with tumor-associated myeloid cells playing vital roles in tumor immune evasion (Fig.2).

4.1.1 Tumor-associated macrophages

Macrophages is known to be classified into two subtypes: M1 and M2 macrophages. M1 macrophages prompt both inflammatory and anti-tumor responses. Activated M2 macrophages secrete a series of inflammatory response factors that participate in regulating the inflammatory microenvironment [120]. TAMs usually express an M2 phenotype and are crucial components of the tumor inflammatory microenvironment [121]. Furthermore, TAMs involve in tumor occurrence, progression and migration, which are pivotal immune cells and associated with liver CSCs formation [122]. This section will discuss the relationship between TAMs and liver CSCs to provide a novel insight for cancer treatment from the perspective of targeting TAMs.

TAMs form a “niche” to maintain the self-renewal of CSCs. Cytokines secreted by macrophages regulate CSCs properties via multiple pathways [123]. For example, TAMs secret IL-6 to activate IL-6/STAT3 axis to promote CD44⁺ HCC cells expansion and liver tumorigenesis [78]. Similarly, CCL17 produced by activated M2 macrophages triggers the activation of TGF-β1 and Wnt/β-catenin signaling in HCC cells, which promotes the stemness and EMT process of HCC cells [124]. TAMs produce TNF-α to promote HCC cell stemness via Wnt/β-catenin axis [79]. Consistently, TAMs also produce TGF-β1 to induce EMT and stemness properties in liver cancer cells, which gain higher metastasis and invasive capability [80]. Moreover, HCC cell-derived TGF-β drives T cell immunoglobulin and mucin domain 3 (TIM-3) expression on TAMs, then TIM-3 stimulates STAT6 and nuclear factor-κB (NF-κB) pathway, which foster TAM to produce IL-10 and stimulate HCC cell migration [125]. Furthermore, IL-8 secreted by macrophages regulates downstream JAK2/STAT3/Snail signaling to trigger the invasion properties of HCC cells [126]. Moreover, TAMs secret CXCL2, CXCL8, and TNF-α to recruit neutrophils into HCC milieus and increase the production of pro-stemness factor oncostatin M (OSM) in neutrophils, finally leading to HCC metastasis and malignant progression [127]. In addition, HCC cells treated with TAM extracellular vesicles or miR-125a/b targeting liver CSC marker CD90 could suppress cell proliferation and stem cell properties. This finding suggests that targeting CD90 has an inhibitory effect on liver CSCs [128]. Consequently, TAM-CSC communication is related with tumor development.

Liver CSCs promote migration and recruit macrophages to tumor sites by secreting chemokines. CSCs possess an immunosuppressive program involved in macrophages recruitment and trigger them toward M2 polarization at the tumor locus, this ability of CSCs is generally found in HCC through activating NF-κB signaling pathways and secreting cytokines such as IL-8 and IL-10 [129]. For instance, CD133⁺ cells secrete IL-8 to induce TAMs M2 polarization in HCC [123]. Similarly, HCC-derived IL-8 attracts more M2-type TAMs to promote a pro-oncogenic inflammatory microenvironment, and aberrant activation of the neurotensin (NTS)/IL-8 pathway stimulates the invasion potential of HCC cells [123]. Notably, CSCs escape immune surveillance by exerting TAMs immunosuppressive functions. CD24 expressed on tumor cells interacts with sialic-acid binding Ig-like lectin 10 (Siglec-10) on TAMs to promote HCC progression [130]. Similarly, CD47 expressed on tumor cells interacts with signal regulatory protein alpha (SIPRα) on macrophages to protect liver CSCs from being phagocytosed [30]. These observations demonstrate that both immunosuppression and stemness phenotypes can be targeted simultaneously between the crosstalk of CSCs and macrophages.

4.1.2 Tumor-associated neutrophils

TANs are classical inflammatory cells associated with tumorigenesis and development in the TME [84]. However, the direct interplay between TANs and HCC cells remains unclear [131]. Recently, it has been demonstrated that BMP2 and TGF-β2 upregulate miR-301b-3p expression in HCC cells, and subsequently block limbic system-associated membrane protein (LSAMP) and CYLD lysine 63 deubiquitinase (CYLD) expression, thus contributing to HCC stemness [84]. In addition, NF-κB signaling is activated in TAN-induced HCC stem cell like cells, higher CXCL5 is generated, and more TANs infiltration is recruited, which forms a positive feedback network. It is also validated in HCC clinical patients [84]. Of note, the interaction of CAFs, HCC cells, and TANs are connected with a network of cytokines. For instance, cardiotrophin-like cytokine factor 1 (CLCF1) derived from CAFs upregulates the expression of CXCL6 and TGF-β in tumor cells, driving TANs N2 polarization and subsequently promoting HCC cells stem-cell like characteristics [132]. In conclusion, all of above observations suggest that TANs modulate liver CSCs and are effective targets for HCC treatment.

4.1.3 Myeloid-derived suppressor cells

MDSCs represent a heterogeneous population of immature myeloid cells. MDSCs could disrupt the effect of DCs and inhibit T cell infiltration [133–135]. Emerging evidence implicates that MDSCs inhibit antitumor immune response in HCC patients [135]. Drug-resistant HCC cells are generally considered to have CSC properties [136]. Drug-resistant HCC cells-derived IL-6 activates MDSCs and facilitates the immunosuppressive function of MDSCs with a higher production of NO and ROS [136]. Furthermore, tumor cell-derived stem cell factor (SCF) drives MDSCs expansion and promotes the proliferation of liver cancer cells [137]. Moreover, HSC-monocytic MDSCs (M-MDSC) crosstalk promotes HCC progression. Activated HSC triggers monocyte-intrinsic p38 MAPK signaling to induce M-MDSCs accumulation and promote HCC development [138]. In addition, IL-1β triggers solute carrier family 7 member 11 (SLC7A11) expression and subsequently upregulates PD-L1 and colony stimulating factor-1 (CSF1) expression via α-ketoglutarate (α-KG)-HIF1α signaling pathway, which promotes TAMs and MDSCs infiltration, and thus contributing to HCC invasion [139]. Interestingly, CAFs secret IL-6 and IL-33 to mediate the activation of 5-lipoxygenase (5-LO) metabolism in MDSCs, 5-LO mediates the function of CD33⁺ MDSCs to stimulate intrahepatic cholangiocarcinoma (ICC) stemness through its downstream leukotriene B4 (LTB4)-leukotriene B4 receptor type 2 (BLT2) axis via the activation of AKT/mTORC1 signaling [140]. Therefore, targeting MDSCs probably provides a novel strategy for improving HCC clinical treatment.

4.2 Antitumorigenic immune cells

Antitumorigenic immune cells in the TME predominantly include tumor-infiltrating lymphocytes (TILs) and DCs infiltrate the TME. The presence of a large number of TILs and DCs indicates that the body has initiated an immune response against the tumor. If the human immune system response to cancer is likened to a war, antitumorigenic immune cells are a “warrior” that rushes to the front line from all over the body to fight against cancer cells [141]. The amount of TILs and DCs in tumors significantly indicates the prognosis and response to immunotherapy in HCC patients (Fig.3) [142].

4.2.1 T lymphocytes

T cells are vital components of immunosurveillance and play an essential role in long-term antigen-specific immune responses. The process of liver CSCs regulating T cell activity is involved in distinct pathways. Liver CSCs can negatively regulate T cell activity through transforming the activity of Treg cells [143]. TGF-β signaling could suppress miR-34a and increase the production of chemokine CCL22 in CD44⁺ liver CSCs, which leads to Treg cells recruitment and thus favoring HCC immune escape [144]. Tumor cells and immune cells within immune-high HCC tumors linked to highly PD-L1 expression [145]. The presence of lymphocytes significantly correlates with HCC patient outcomes, PD-L1 expression is related to the HCC progenitor properties (expressed CK19 and SALL4) in an analysis of 217 HCC clinical patients [145,146], implying the efficacy of anti-PD-1 immunotherapy directly acting on CD8⁺ T cells when it is applied for HCC treatment. Additionally, β-catenin/STT3 signaling contributes to PD-L1 enrichment through modulating glycosylation and stabilizing PD-L1 in CSCs, which is crucial for CSC immune evasion [147]. Moreover, IL-6 activates JAK1 signaling and JAK1 phosphorylates PD-L1 to catalyze PD-L1 glycosylation and stabilization, which is also observed in HCC patient samples [148]. Of note, nicotinamide adenine dinucleotide (NAD⁺) metabolism is necessary for the maintenance of liver CSCs. MRPS5 increases the proportion of NAD⁺, which is essential for enhancing the function of mitochondria in liver CSCs [149]. Recently, we have demonstrated that NAD⁺ metabolism maintains PD-L1 expression on HCC cells in response to IFN-γ, which is derived from T cells. In turn, PD-L1 expression on HCC cells interacts with PD-1 on T cells to escape T cell-mediated immune surveillance [94]. In addition, Trp53^−/− HCC cells activate β-catenin signaling pathway to escape immune surveillance by impairing T cell cytotoxic activity, indicating that β-catenin reduces the DCs recruitment and subsequently downregulates T cell activity and thus contributing to tumor progression [150]. β-catenin activation could influence HCC immune microenvironment and promote HCC progression. Heterogeneous nuclear ribonucleoprotein M (HNRNPM) deficiency inhibits Wnt/β-catenin signaling and contributes to CD8⁺ T cell infiltration in HCC, thus increasing the efficiency of anti-PD-1 immunotherapy. HNRNPM upregulates FZD3 expression and activates Wnt/β-catenin signaling by regulating the alternative splicing (AS) of MBD2, FZD3 and β-catenin further regulate stemness by targeting CSC marker OCT4 and SOX2 [151]. Separately, the hyaluronan synthesis inhibitor 4-methylumbelliferone (4mu) reduces the proportion of CD47⁺ liver CSCs and elicits a T cell response induced by IL-12 [152]. In conclusion, the cytotoxic activity of tumor infiltrates contributes to immune surveillance by inhibiting liver CSC stemness properties. HCC forms a robust immune tolerance microenvironment because HCC patients have been in a state of low immune response for a long time and tended to develop resistance to tumor immunotherapy. Therefore, it has important clinical and therapeutic significance to study how to improve HCC immune resistance and enhance T cell-mediated immune response.

4.2.2 Natural killer cells

The NK cell population is an important subpopulation of cytotoxic lymphocytes in HCC progression. It is known that GEP and ABCB5 expression in liver CSCs contribute to HCC poor outcomes [153]. GEP reduces MHC class I chain-related molecule A (MICA) expression, ligand for NK stimulatory receptor NK group 2 member D (NKG2D), and upregulates human leukocyte antigen-E (HLA-E), ligand for NK inhibitory receptor CD94/NKG2A on NK cells. Thus, GEP-expressing CSCs resistant NK cell-mediated cytotoxicity and promote HCC tumor immune escape [154]. Additionally, carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is upregulated on EpCAM-high liver CSCs, it contributes to resistance to NK cell cytotoxicity and promotes HCC development, which indicates that homophilic interaction of CEACAM1 is critical for alleviative NK cell cytotoxic effect on CEACAM1^high liver CSCs [155]. Moreover, HCC cell-derived exosomal circular ubiquitin-like with PHD and circUHRF1 inhibits NK cell-secreted IFN-γ and TNF-α. circUHRF1 suppresses NK cell-mediated effect by increasing TIM-3 expression and downregulating miR-449c-5p in HCC [156]. In summary, NK cell-mediated killing activities are regulated by liver CSCs through some critical mediators, which further suggest that NK cell-mediated immune response is vital for HCC progression.

4.2.3 Dendritic cells

DCs are antigen-presenting cells that are closely related to immune responses and have a vital role in exerting the T cell-mediated antitumor immune response [157]. A recently research has suggested that DCs and cytokine-induced killer cells (CIKs) upregulate caspase-3 expression and decrease proliferating cell nuclear antigen (PCNA) expression to increase the apoptosis ratio of liver CSCs [89]. Arf1-mediated lipid metabolism maintains CSCs. Arf1 ablation in CSCs results in metabolic stress including mitochondrial defects and endoplasmic reticulum (ER) stress, releasing damage-associated molecular patterns (DAMPs), which potentiate anti-tumor immune response through DCs activation and T cell infiltration enhancement [90]. Accumulating evidence has demonstrated that liver CSCs contribute to the evasion of immunosurveillance by altering the phenotype of DCs and impairing their recruitment [91,92]. RIG-1 deficiency leads to accumulation of p-STAT3, and p-STAT3 could induce TGF-β1 expression. RIG-I-deficient HCC cells with stemness-associated characteristics alter the roles of DCs from immunostimulatory to immunosuppressive through upregulating TGF-β1 [91]. Moreover, RIG-I deficiency causes more TGF-β1 production that triggers Smad2 and Akt phosphorylation and facilitates the interaction of Smad2/p-Smad2 and Akt/p-Akt in HCC cells, thus promoting HCC stemness [91]. Additionally, β-catenin activation facilitates HCC against anti-PD-1 therapy, which is associated with defective recruitment of DCs [150]. Through the summary of the recent findings about CSCs regulate DCs recruitment, there still remains a need to figure out the association between DCs and liver CSCs.

5 Liver CSCs-targeted immunotherapy

Due to the lack of specific therapeutic targets, HCC has a high mortality rate [158]. Liver CSCs-targeted immunotherapy could be promising therapeutic strategies for HCC [95]. With the emergence of immunotherapy, exciting developments have been made that utilize CSC-specific antigens to harness the immune system’s power and increase the efficacy of HCC treatment. Approaches with direct inhibitory effects include monoclonal antibodies targeting CSC-related cell surface markers, cytotoxic T lymphocyte (CTL)-based immunotherapy, and the use of DC vaccines.

5.1 Antibody-based immunotherapy

Several monoclonal antibodies with direct inhibitory effects have been developed to target CSC-related cell surface markers. The basis of antibody-mediated therapy for targeting CSCs are mainly separated into two segments: antibody-dependent cellular cytotoxicity (ADCC) and the direct inhibitory effect of mAbs [159]. CSC-targeted immunotherapy has been attracted more attention. For instance, EpCAM is recognized as a liver CSC marker and a promising therapeutic target in HCC [160]. An EpCAM/CD3 bispecific antibody (anti-EpCAM bispecific T cell engager (BiTE) 1H8/CD3) recognizing EpCAM can effectively target and eliminate liver CSCs in vitro and in vivo [161]. Additionally, cytokine-induced killer (CIK) cells bound with anti-CD3/anti-CD133 bispecific antibodies induced strong elimination of HCC cells in vitro and in vivo [162]. Other liver CSC marker-specific therapies, such as CD47, CD44, and CD24, related mAbs significantly inhibit HCC tumor progression. CD47 blockade inhibits tumor progression in HCC xenograft models, indicating that targeting CD47 may provide a better prognosis for HCC treatment [163]. CD44 antibody-targeted liposomal nanoparticles effectively target CD44⁺ liver CSCs in preclinical models [164]. Meanwhile, a high-affinity humanized anti-CD24 antibody (hG7-BM3-VcMMAE conjugate) is designed for HCC treatment in vivo [165]. However, it is necessary to validate the clinical efficacy of these liver CSC marker-specific, antibody-based therapies.

5.2 Chimeric antigen receptor-T cells

CTL-based immunotherapy has broad application prospects [166–168]. Alternative methods of generating CSC-specific T cells include genetically engineered chimeric antigen receptor (CAR)-T cells. Indeed, CAR-T cells could recognize specific antigens on CSCs to achieve their targeted effects [169]. Based on the liver CSC markers, several CAR-T-related clinical trials are ongoing. Various CD133⁺ late-stage metastatic malignancies, including HCC, have been treated with CD133-directed CAR-T cells in phase I clinical trials published in 2018 [170]. Moreover, a phase I clinical study (NCT02541370) using CAR-CD133 T cells to treat patients with advanced and CD133⁺ tumors, including 14 HCC patients, results show that 1 HCC patient achieve partial remission, 9 HCC patients acquire stable disease, implying that CD133-directed CAR-T cell infusion has antitumor activity in advanced HCC [170]. Similarly, a phase II clinical trial (NCT02541370) verified that CAR-CD133 T cells is feasible and have controllable toxicities in HCC [171]. Moreover, DLK1-directed CAR-T cells effectively eliminate DLK1-positive HCC cells in vitro and in vivo, indicating that DLK1 also could act as a therapeutic target for HCC treatment [172]. Additionally, one CAR-EpCAM T cell clinical trial (NCT03013712) has been organized and is recruiting EpCAM-positive cancer (including HCC), the efficacy is yet to be determined. Collectively, using CAR-T cells to target CSCs is a promising approach for HCC treatment [173]. Although these results are promising, one hindrance in the effective use of CAR-T therapy is toxicity-related challenges (the reduced levels of hemoglobin and platelets) [174]. The other challenge is antigen selection for CAR designing, as tumor antigens are also expressed on the surface of non-cancer cells, leading to target miss-effect and off-target effects, thus hindering therapeutic efficacy [175].

5.3 Dendritic cell vaccines

DC vaccines facilitate the immune system’s recognition and eradication of tumor cells. Immune response mediated by DCs and triggered by tumor-specific antigens is generated using CSC-derived materials. For instance, pulsing DCs with CD44 and EpCAM peptides facilitate the production of mature DCs, which stimulate T cell activity and produce effective CTLs [176]. In addition, ICT-121, a dendritic cell vaccine targeted at CD133, is shown to promote cytolytic T cell responses against CD133⁺ CSCs in a phase I trial (NCT02049489) [177]. Moreover, ANXA3-transfected DCs could also potently stimulate the killing activity of liver CSCs, indicating that ANXA3 maybe a novel target for CSC-targeted immunotherapy [178]. Additionally, a new work focus on increasing the efficacy of DC-based HCC neoantigen nano-vaccine by remodeling TANs [179].

6 Conclusions

Liver CSCs are a fraction of HCC cells with stem cell characteristics account for HCC initiation and therapeutic resistance. Much effort has been made to identify liver CSC markers and altered signaling pathways during recent decades. In this perspective, these discoveries are essential for improving HCC clinical treatment. To achieve stable remission and even cure aggressive malignancies, it is imperative to eliminate CSCs and block their signaling pathways. Remarkably, we illustrate the interplay of CSCs with individual immune cell lineages, and the mechanisms of crosstalk between them. These findings will aid in the development of novel strategies for effectively targeting CSCs. Liver CSCs-specific therapies require to be further investigated for clinical application. Nevertheless, some important issues remain unresolved. For instance, some of the crucial markers of liver CSCs are also shared by normal stem cells. Molecular differences between CSCs and normal stem cells need to be identified in order to target them specifically without implicating normal stem cells. Ultimately, targeting CSCs specifically increases the efficacy of existing therapies and provides a framework for the development of new therapeutic regimens with long-term clinical benefits. It is expected to identify more effective liver CSC markers and develop more specific anti-CSC marker therapies in the future.

References

[1] Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, Bray F. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 2021; 71(3): 209–249

[2] Huang A, Yang XR, Chung WY, Dennison AR, Zhou J. Targeted therapy for hepatocellular carcinoma. Signal Transduct Target Ther 2020; 5(1): 146

[3] Jörs S, Jeliazkova P, Ringelhan M, Thalhammer J, Dürl S, Ferrer J, Sander M, Heikenwalder M, Schmid RM, Siveke JT, Geisler F. Lineage fate of ductular reactions in liver injury and carcinogenesis. J Clin Invest 2015; 125(6): 2445–2457

[4] Llovet JM, Bruix J. Systematic review of randomized trials for unresectable hepatocellular carcinoma: chemoembolization improves survival. Hepatology 2003; 37(2): 429–442

[5] Oishi N, Yamashita T, Kaneko S. Molecular biology of liver cancer stem cells. Liver Cancer 2014; 3(2): 71–84

[6] Joo I, Kim H, Lee JM. Cancer stem cells in primary liver cancers: pathological concepts and imaging findings. Korean J Radiol 2015; 16(1): 50–68

[7] Hinshaw DC, Shevde LA. The tumor microenvironment innately modulates cancer progression. Cancer Res 2019; 79(18): 4557–4566

[8] Dong KS, Chen Y, Yang G, Liao ZB, Zhang HW, Liang HF, Chen XP, Dong HH. TGF-β1 accelerates the hepatitis B virus X-induced malignant transformation of hepatic progenitor cells by upregulating miR-199a-3p. Oncogene 2020; 39(8): 1807–1820

[9] Bayik D, Lathia JD. Cancer stem cell-immune cell crosstalk in tumour progression. Nat Rev Cancer 2021; 21(8): 526–536

[10] He G, Dhar D, Nakagawa H, Font-Burgada J, Ogata H, Jiang Y, Shalapour S, Seki E, Yost SE, Jepsen K, Frazer KA, Harismendy O, Hatziapostolou M, Iliopoulos D, Suetsugu A, Hoffman RM, Tateishi R, Koike K, Karin M. Identification of liver cancer progenitors whose malignant progression depends on autocrine IL-6 signaling. Cell 2013; 155(2): 384–396

[11] Wu K, Ding J, Chen C, Sun W, Ning BF, Wen W, Huang L, Han T, Yang W, Wang C, Li Z, Wu MC, Feng GS, Xie WF, Wang HY. Hepatic transforming growth factor beta gives rise to tumor-initiating cells and promotes liver cancer development. Hepatology 2012; 56(6): 2255–2267

[12] Tang Y, Kitisin K, Jogunoori W, Li C, Deng CX, Mueller SC, Ressom HW, Rashid A, He AR, Mendelson JS, Jessup JM, Shetty K, Zasloff M, Mishra B, Reddy EP, Johnson L, Mishra L. Progenitor/stem cells give rise to liver cancer due to aberrant TGF-β and IL-6 signaling. Proc Natl Acad Sci USA 2008; 105(7): 2445–2450

[13] Jing Y, Sun K, Liu W, Sheng D, Zhao S, Gao L, Wei L. Tumor necrosis factor-α promotes hepatocellular carcinogenesis through the activation of hepatic progenitor cells. Cancer Lett 2018; 434: 22–32

[14] Holczbauer Á, Factor VM, Andersen JB, Marquardt JU, Kleiner DE, Raggi C, Kitade M, Seo D, Akita H, Durkin ME, Thorgeirsson SS. Modeling pathogenesis of primary liver cancer in lineage-specific mouse cell types. Gastroenterology 2013; 145(1): 221–231

[15] Mu X, Español-Suñer R, Mederacke I, Affò S, Manco R, Sempoux C, Lemaigre FP, Adili A, Yuan D, Weber A, Unger K, Heikenwälder M, Leclercq IA, Schwabe RF. Hepatocellular carcinoma originates from hepatocytes and not from the progenitor/biliary compartment. J Clin Invest 2015; 125(10): 3891–3903

[16] Huch M, Dollé L. The plastic cellular states of liver cells: are EpCAM and Lgr5 fit for purpose?. Hepatology 2016; 64(2): 652–662

[17] Chaffer CL, Brueckmann I, Scheel C, Kaestli AJ, Wiggins PA, Rodrigues LO, Brooks M, Reinhardt F, Su Y, Polyak K, Arendt LM, Kuperwasser C, Bierie B, Weinberg RA. Normal and neoplastic nonstem cells can spontaneously convert to a stem-like state. Proc Natl Acad Sci USA 2011; 108(19): 7950–7955

[18] Fekir K, Dubois-Pot-Schneider H, Désert R, Daniel Y, Glaise D, Rauch C, Morel F, Fromenty B, Musso O, Cabillic F, Corlu A. Retrodifferentiation of human tumor hepatocytes to stem cells leads to metabolic reprogramming and chemoresistance. Cancer Res 2019; 79(8): 1869–1883

[19] Li XF, Chen C, Xiang DM, Qu L, Sun W, Lu XY, Zhou TF, Chen SZ, Ning BF, Cheng Z, Xia MY, Shen WF, Yang W, Wen W, Lee TKW, Cong WM, Wang HY, Ding J. Chronic inflammation-elicited liver progenitor cell conversion to liver cancer stem cell with clinical significance. Hepatology 2017; 66(6): 1934–1951

[20] Matsumoto T, Takai A, Eso Y, Kinoshita K, Manabe T, Seno H, Chiba T, Marusawa H. Proliferating EpCAM-positive ductal cells in the inflamed liver give rise to hepatocellular carcinoma. Cancer Res 2017; 77(22): 6131–6143

[21] Ang CH, Hsu SH, Guo F, Tan CT, Yu VC, Visvader JE, Chow PKH, Fu NY. Lgr5⁺ pericentral hepatocytes are self-maintained in normal liver regeneration and susceptible to hepatocarcinogenesis. Proc Natl Acad Sci USA 2019; 116(39): 19530–19540

[22] Tsui YM, Chan LK, Ng IO. Cancer stemness in hepatocellular carcinoma: mechanisms and translational potential. Br J Cancer 2020; 122(10): 1428–1440

[23] Lee TK, Guan XY, Ma S. Cancer stem cells in hepatocellular carcinoma — from origin to clinical implications. Nat Rev Gastroenterol Hepatol 2022; 19(1): 26–44

[24] Mishra L, Banker T, Murray J, Byers S, Thenappan A, He AR, Shetty K, Johnson L, Reddy EP. Liver stem cells and hepatocellular carcinoma. Hepatology 2009; 49(1): 318–329

[25] Jang JW, Song Y, Kim SH, Kim JS, Kim KM, Choi EK, Kim J, Seo HR. CD133 confers cancer stem-like cell properties by stabilizing EGFR-AKT signaling in hepatocellular carcinoma. Cancer Lett 2017; 389: 1–10

[26] Yang ZF, Ho DW, Ng MN, Lau CK, Yu WC, Ngai P, Chu PWK, Lam CT, Poon RTP, Fan ST. Significance of CD90⁺ cancer stem cells in human liver cancer. Cancer Cell 2008; 13(2): 153–166

[27] Mima K, Okabe H, Ishimoto T, Hayashi H, Nakagawa S, Kuroki H, Watanabe M, Beppu T, Tamada M, Nagano O, Saya H, Baba H. CD44s regulates the TGF-β-mediated mesenchymal phenotype and is associated with poor prognosis in patients with hepatocellular carcinoma. Cancer Res 2012; 72(13): 3414–3423

[28] Yamashita T, Forgues M, Wang W, Kim JW, Ye Q, Jia H, Budhu A, Zanetti KA, Chen Y, Qin LX, Tang ZY, Wang XW. EpCAM and α-fetoprotein expression defines novel prognostic subtypes of hepatocellular carcinoma. Cancer Res 2008; 68(5): 1451–1461

[29] Haraguchi N, Ishii H, Mimori K, Tanaka F, Ohkuma M, Kim HM, Akita H, Takiuchi D, Hatano H, Nagano H, Barnard GF, Doki Y, Mori M. CD13 is a therapeutic target in human liver cancer stem cells. J Clin Invest 2010; 120(9): 3326–3339

[30] Lee TKW, Cheung VCH, Lu P, Lau EYT, Ma S, Tang KH, Tong M, Lo J, Ng IOL. Blockade of CD47-mediated cathepsin S/protease-activated receptor 2 signaling provides a therapeutic target for hepatocellular carcinoma. Hepatology 2014; 60(1): 179–191

[31] Xu X, Liu RF, Zhang X, Huang LY, Chen F, Fei QL, Han ZG. DLK1 as a potential target against cancer stem/progenitor cells of hepatocellular carcinoma. Mol Cancer Ther 2012; 11(3): 629–638

[32] Liu S, Li N, Yu X, Xiao X, Cheng K, Hu J, Wang J, Zhang D, Cheng S, Liu S. Expression of intercellular adhesion molecule 1 by hepatocellular carcinoma stem cells and circulating tumor cells. Gastroenterology 2013; 144(5): 1031–1041.e10

[33] Zhao W, Wang L, Han H, Jin K, Lin N, Guo T, Chen Y, Cheng H, Lu F, Fang W, Wang Y, Xing B, Zhang Z. 1B50-1, a mAb raised against recurrent tumor cells, targets liver tumor-initiating cells by binding to the calcium channel α2δ1 subunit. Cancer Cell 2013; 23(4): 541–556

[34] Kawai T, Yasuchika K, Ishii T, Katayama H, Yoshitoshi EY, Ogiso S, Kita S, Yasuda K, Fukumitsu K, Mizumoto M, Hatano E, Uemoto S. Keratin 19, a cancer stem cell marker in human hepatocellular carcinoma. Clin Cancer Res 2015; 21(13): 3081–3091

[35] Tang KH, Ma S, Lee TK, Chan YP, Kwan PS, Tong CM, Ng IO, Man K, To KF, Lai PB, Lo CM, Guan XY, Chan KW. CD133⁺ liver tumor-initiating cells promote tumor angiogenesis, growth, and self-renewal through neurotensin/interleukin-8/CXCL1 signaling. Hepatology 2012; 55(3): 807–820

[36] Hur W, Ryu JY, Kim HU, Hong SW, Lee EB, Lee SY, Yoon SK. Systems approach to characterize the metabolism of liver cancer stem cells expressing CD133. Sci Rep 2017; 7(1): 45557

[37] Yang ZF, Ngai P, Ho DW, Yu WC, Ng MNP, Lau CK, Li MLY, Tam KH, Lam CT, Poon RTP, Fan ST. Identification of local and circulating cancer stem cells in human liver cancer. Hepatology 2008; 47(3): 919–928

[38] Ishimoto T, Nagano O, Yae T, Tamada M, Motohara T, Oshima H, Oshima M, Ikeda T, Asaba R, Yagi H, Masuko T, Shimizu T, Ishikawa T, Kai K, Takahashi E, Imamura Y, Baba Y, Ohmura M, Suematsu M, Baba H, Saya H. CD44 variant regulates redox status in cancer cells by stabilizing the xCT subunit of system xc^- and thereby promotes tumor growth. Cancer Cell 2011; 19(3): 387–400

[39] Kim JW, Ye Q, Forgues M, Chen Y, Budhu A, Sime J, Hofseth LJ, Kaul R, Wang XW. Cancer-associated molecular signature in the tissue samples of patients with cirrhosis. Hepatology 2004; 39(2): 518–527

[40] Kawai T, Yasuchika K, Seo S, Higashi T, Ishii T, Miyauchi Y, Kojima H, Yamaoka R, Katayama H, Yoshitoshi EY, Ogiso S, Kita S, Yasuda K, Fukumitsu K, Nakamoto Y, Hatano E, Uemoto S. Identification of keratin 19-positive cancer stem cells associating human hepatocellular carcinoma using ¹⁸F-fluorodeoxyglucose positron emission tomography. Clin Cancer Res 2017; 23(6): 1450–1460

[41] Wang N, Li MY, Liu Y, Yu J, Ren J, Zheng Z, Wang S, Yang S, Yang SL, Liu LP, Hu BG, Chong CC, Merchant JL, Lai PB, Chen GG. ZBP-89 negatively regulates self-renewal of liver cancer stem cells via suppression of Notch1 signaling pathway. Cancer Lett 2020; 472: 70–80

[42] Wang N, Wang S, Yang SL, Liu LP, Li MY, Lai PBS, Chen GG. Targeting ZBP-89 for the treatment of hepatocellular carcinoma. Expert Opin Ther Targets 2018; 22(10): 817–822

[43] Nio K, Yamashita T, Kaneko S. The evolving concept of liver cancer stem cells. Mol Cancer 2017; 16(1): 4

[44] Monga SPS. Role of Wnt/β-catenin signaling in liver metabolism and cancer. Int J Biochem Cell Biol 2011; 43(7): 1021–1029

[45] Chang PH, Sekine K, Chao HM, Hsu SH, Chern E. Chitosan promotes cancer progression and stem cell properties in association with Wnt signaling in colon and hepatocellular carcinoma cells. Sci Rep 2017; 8(1): 45751

[46] Fan Z, Duan J, Wang L, Xiao S, Li L, Yan X, Yao W, Wu L, Zhang S, Zhang Y, Li Y, Zhu X, Hu Y, Zhang D, Jiao S, Xu X. PTK2 promotes cancer stem cell traits in hepatocellular carcinoma by activating Wnt/β-catenin signaling. Cancer Lett 2019; 450: 132–143

[47] Ram Makena M, Gatla H, Verlekar D, Sukhavasi SK, Pandey MC, Pramanik K. Wnt/β-catenin signaling: the culprit in pancreatic carcinogenesis and therapeutic resistance. Int J Mol Sci 2019; 20(17): 4242

[48] Yamashita T, Ji J, Budhu A, Forgues M, Yang W, Wang HY, Jia H, Ye Q, Qin LX, Wauthier E, Reid LM, Minato H, Honda M, Kaneko S, Tang ZY, Wang XW. EpCAM-positive hepatocellular carcinoma cells are tumor-initiating cells with stem/progenitor cell features. Gastroenterology 2009; 136(3): 1012–1024

[49] Ma S, Chan KW, Hu L, Lee TK, Wo JY, Ng IO, Zheng BJ, Guan XY. Identification and characterization of tumorigenic liver cancer stem/progenitor cells. Gastroenterology 2007; 132(7): 2542–2556

[50] Zhu J, Yu H, Chen S, Yang P, Dong Z, Ling Y, Tang H, Bai S, Yang W, Tang L, Shen F, Wang H, Wen W. Prognostic significance of combining high mobility group Box-1 and OV-6 expression in hepatocellular carcinoma. Sci China Life Sci 2018; 61(8): 912–923

[51] Mokkapati S, Niopek K, Huang L, Cunniff KJ, Ruteshouser EC, deCaestecker M, Finegold MJ, Huff V. β-catenin activation in a novel liver progenitor cell type is sufficient to cause hepatocellular carcinoma and hepatoblastoma. Cancer Res 2014; 74(16): 4515–4525

[52] Xiang D, Cheng Z, Liu H, Wang X, Han T, Sun W, Li X, Yang W, Chen C, Xia M, Liu N, Yin S, Jin G, Lee T, Dong L, Hu H, Wang H, Ding J. Shp2 promotes liver cancer stem cell expansion by augmenting β-catenin signaling and predicts chemotherapeutic response of patients. Hepatology 2017; 65(5): 1566–1580

[53] Leung HW, Leung CON, Lau EY, Chung KPS, Mok EH, Lei MML, Leung RWH, Tong M, Keng VW, Ma C, Zhao Q, Ng IOL, Ma S, Lee TK. EPHB2 activates β-catenin to enhance cancer stem cell properties and drive sorafenib resistance in hepatocellular carcinoma. Cancer Res 2021; 81(12): 3229–3240

[54] Yamashita T, Budhu A, Forgues M, Wang XW. Activation of hepatic stem cell marker EpCAM by Wnt-beta-catenin signaling in hepatocellular carcinoma. Cancer Res 2007; 67(22): 10831–10839

[55] Kahraman DC, Kahraman T, Cetin-Atalay R. Targeting PI3K/Akt/mTOR pathway identifies differential expression and functional role of IL8 in liver cancer stem cell enrichment. Mol Cancer Ther 2019; 18(11): 2146–2157

[56] Wang XQ, Zhang W, Lui ELH, Zhu Y, Lu P, Yu X, Sun J, Yang S, Poon RTP, Fan ST. Notch1-Snail1-E-cadherin pathway in metastatic hepatocellular carcinoma. Int J Cancer 2012; 131(3): E163–E172

[57] Xiao S, Chang RM, Yang MY, Lei X, Liu X, Gao WB, Xiao JL, Yang LY. Actin-like 6A predicts poor prognosis of hepatocellular carcinoma and promotes metastasis and epithelial-mesenchymal transition. Hepatology 2016; 63(4): 1256–1271

[58] Zhu P, Wang Y, Du Y, He L, Huang G, Zhang G, Yan X, Fan Z. C8orf4 negatively regulates self-renewal of liver cancer stem cells via suppression of NOTCH2 signalling. Nat Commun 2015; 6(1): 7122

[59] Wang R, Li Y, Tsung A, Huang H, Du Q, Yang M, Deng M, Xiong S, Wang X, Zhang L, Geller DA, Cheng B, Billiar TR. iNOS promotes CD24⁺CD133⁺ liver cancer stem cell phenotype through a TACE/ADAM17-dependent Notch signaling pathway. Proc Natl Acad Sci USA 2018; 115(43): E10127–E10136

[60] Lin L, Amin R, Gallicano GI, Glasgow E, Jogunoori W, Jessup JM, Zasloff M, Marshall JL, Shetty K, Johnson L, Mishra L, He AR. The STAT3 inhibitor NSC 74859 is effective in hepatocellular cancers with disrupted TGF-β signaling. Oncogene 2009; 28(7): 961–972

[61] Wang C, Yang W, Yan HX, Luo T, Zhang J, Tang L, Wu FQ, Zhang HL, Yu LX, Zheng LY, Li YQ, Dong W, He YQ, Liu Q, Zou SS, Lin Y, Hu L, Li Z, Wu MC, Wang HY. Hepatitis B virus X (HBx) induces tumorigenicity of hepatic progenitor cells in 3,5-diethoxycarbonyl-1,4-dihydrocollidine-treated HBx transgenic mice. Hepatology 2012; 55(1): 108–120

[62] Lee TK, Castilho A, Cheung VC, Tang KH, Ma S, Ng IO. CD24⁺ liver tumor-initiating cells drive self-renewal and tumor initiation through STAT3-mediated NANOG regulation. Cell Stem Cell 2011; 9(1): 50–63

[63] Xie C, Zhu J, Wang X, Chen J, Geng S, Wu J, Zhong C, Li X. Tobacco smoke induced hepatic cancer stem cell-like properties through IL-33/p38 pathway. J Exp Clin Cancer Res 2019; 38(1): 39

[64] Toh TB, Lim JJ, Hooi L, Rashid MBMA, Chow EK. Targeting Jak/Stat pathway as a therapeutic strategy against SP/CD44⁺ tumorigenic cells in Akt/β-catenin-driven hepatocellular carcinoma. J Hepatol 2020; 72(1): 104–118

[65] Schiavoni G, Gabriele L, Mattei F. The tumor microenvironment: a pitch for multiple players. Front Oncol 2013; 3: 90

[66] Fang T, Lv H, Lv G, Li T, Wang C, Han Q, Yu L, Su B, Guo L, Huang S, Cao D, Tang L, Tang S, Wu M, Yang W, Wang H. Tumor-derived exosomal miR-1247-3p induces cancer-associated fibroblast activation to foster lung metastasis of liver cancer. Nat Commun 2018; 9(1): 191

[67] Tian B, Luo Q, Ju Y, Song G. A soft matrix enhances the cancer stem cell phenotype of HCC cells. Int J Mol Sci 2019; 20(11): 2831

[68] Loong JH, Wong TL, Tong M, Sharma R, Zhou L, Ng KY, Yu HJ, Li CH, Man K, Lo CM, Guan XY, Lee TK, Yun JP, Ma SK. Glucose deprivation-induced aberrant FUT1-mediated fucosylation drives cancer stemness in hepatocellular carcinoma. J Clin Invest 2021; 131(11): e143377

[69] Liu Z, Dai X, Wang T, Zhang C, Zhang W, Zhang W, Zhang Q, Wu K, Liu F, Liu Y, Wu J. Hepatitis B virus PreS1 facilitates hepatocellular carcinoma development by promoting appearance and self-renewal of liver cancer stem cells. Cancer Lett 2017; 400: 149–160

[70] Ding WB, Wang MC, Yu J, Huang G, Sun DP, Liu L, Zhang JN, Yang Y, Liu H, Zhou WP, Yang F, Yuan SX. HBV/pregenomic RNA increases the stemness and promotes the development of HBV-related HCC through reciprocal regulation with insulin-like growth factor 2 mRNA-binding protein 3. Hepatology 2021; 74(3): 1480–1495

[71] Lau EYT, Lo J, Cheng BYL, Ma MKF, Lee JMF, Ng JKY, Chai S, Lin CH, Tsang SY, Ma S, Ng IOL, Lee TKW. Cancer-associated fibroblasts regulate tumor-initiating cell plasticity in hepatocellular carcinoma through c-Met/FRA1/HEY1 signaling. Cell Rep 2016; 15(6): 1175–1189

[72] Liu C, Liu L, Chen X, Cheng J, Zhang H, Zhang C, Shan J, Shen J, Qian C. LSD1 stimulates cancer-associated fibroblasts to drive notch3-dependent self-renewal of liver cancer stem-like cells. Cancer Res 2018; 78(4): 938–949

[73] Xiong S, Wang R, Chen Q, Luo J, Wang J, Zhao Z, Li Y, Wang Y, Wang X, Cheng B. Cancer-associated fibroblasts promote stem cell-like properties of hepatocellular carcinoma cells through IL-6/STAT3/Notch signaling. Am J Cancer Res 2018; 8(2): 302–316

[74] Tavora B, Mederer T, Wessel KJ, Ruffing S, Sadjadi M, Missmahl M, Ostendorf BN, Liu X, Kim JY, Olsen O, Welm AL, Goodarzi H, Tavazoie SF. Tumoural activation of TLR3-SLIT2 axis in endothelium drives metastasis. Nature 2020; 586(7828): 299–304

[75] Conigliaro A, Costa V, Lo Dico A, Saieva L, Buccheri S, Dieli F, Manno M, Raccosta S, Mancone C, Tripodi M, De Leo G, Alessandro R. CD90⁺ liver cancer cells modulate endothelial cell phenotype through the release of exosomes containing H19 lncRNA. Mol Cancer 2015; 14(1): 155

[76] Yao H, Liu N, Lin MC, Zheng J. Positive feedback loop between cancer stem cells and angiogenesis in hepatocellular carcinoma. Cancer Lett 2016; 379(2): 213–219

[77] Bishayee A. The role of inflammation and liver cancer. Adv Exp Med Biol 2014; 816: 401–435

[78] Wan S, Zhao E, Kryczek I, Vatan L, Sadovskaya A, Ludema G, Simeone DM, Zou W, Welling TH. Tumor-associated macrophages produce interleukin 6 and signal via STAT3 to promote expansion of human hepatocellular carcinoma stem cells. Gastroenterology 2014; 147(6): 1393–1404

[79] Chen Y, Wen H, Zhou C, Su Q, Lin Y, Xie Y, Huang Y, Qiu Q, Lin J, Huang X, Tan W, Min C, Wang C. TNF-α derived from M2 tumor-associated macrophages promotes epithelial-mesenchymal transition and cancer stemness through the Wnt/β-catenin pathway in SMMC-7721 hepatocellular carcinoma cells. Exp Cell Res 2019; 378(1): 41–50

[80] Fan QM, Jing YY, Yu GF, Kou XR, Ye F, Gao L, Li R, Zhao QD, Yang Y, Lu ZH, Wei LX. Tumor-associated macrophages promote cancer stem cell-like properties via transforming growth factor-beta1-induced epithelial-mesenchymal transition in hepatocellular carcinoma. Cancer Lett 2014; 352(2): 160–168

[81] Liu YC, Yeh CT, Lin KH. Cancer stem cell functions in hepatocellular carcinoma and comprehensive therapeutic strategies. Cells 2020; 9(6): 1331

[82] Chen J, Zheng DX, Yu XJ, Sun HW, Xu YT, Zhang YJ, Xu J. Macrophages induce CD47 upregulation via IL-6 and correlate with poor survival in hepatocellular carcinoma patients. OncoImmunology 2019; 8(11): e1652540

[83] Ye LY, Chen W, Bai XL, Xu XY, Zhang Q, Xia XF, Sun X, Li GG, Hu QD, Fu QH, Liang TB. Hypoxia-induced epithelial-to-mesenchymal transition in hepatocellular carcinoma induces an immunosuppressive tumor microenvironment to promote metastasis. Cancer Res 2016; 76(4): 818–830

[84] Zhou SL, Yin D, Hu ZQ, Luo CB, Zhou ZJ, Xin HY, Yang XR, Shi YH, Wang Z, Huang XW, Cao Y, Fan J, Zhou J. A positive feedback loop between cancer stem-like cells and tumor-associated neutrophils controls hepatocellular carcinoma progression. Hepatology 2019; 70(4): 1214–1230

[85] Schramm C, Oo YH, Lohse AW. Tolerance and autoimmunity in the liver. Semin Immunopathol 2022; 44(4): 393–395

[86] Chaudhary B, Elkord E. Regulatory T cells in the tumor microenvironment and cancer progression: role and therapeutic targeting. Vaccines (Basel) 2016; 4(3): 28

[87] Shi C, Chen Y, Chen Y, Yang Y, Bing W, Qi J. CD4⁺ CD25⁺ regulatory T cells promote hepatocellular carcinoma invasion via TGF-β1-induced epithelial-mesenchymal transition. OncoTargets Ther 2018; 12: 279–289

[88] Rahma OE, Hodi FS. The intersection between tumor angiogenesis and immune suppression. Clin Cancer Res 2019; 25(18): 5449–5457

[89] Yang T, Zhang W, Wang L, Xiao C, Wang L, Gong Y, Huang D, Guo B, Li Q, Xiang Y, Nan Y. Co-culture of dendritic cells and cytokine-induced killer cells effectively suppresses liver cancer stem cell growth by inhibiting pathways in the immune system. BMC Cancer 2018; 18(1): 984

[90] Wang G, Xu J, Zhao J, Yin W, Liu D, Chen W, Hou SX. Arf1-mediated lipid metabolism sustains cancer cells and its ablation induces anti-tumor immune responses in mice. Nat Commun 2020; 11(1): 220

[91] Zhong M, Zhong C, Cui W, Wang G, Zheng G, Li L, Zhang J, Ren R, Gao H, Wang T, Li X, Che J, Gohda E. Induction of tolerogenic dendritic cells by activated TGF-β/Akt/Smad2 signaling in RIG-I-deficient stemness-high human liver cancer cells. BMC Cancer 2019; 19(1): 439

[92] Pardee AD, Shi J, Butterfield LH. Tumor-derived α-fetoprotein impairs the differentiation and T cell stimulatory activity of human dendritic cells. J Immunol 2014; 193(11): 5723–5732

[93] Shankaran V, Ikeda H, Bruce AT, White JM, Swanson PE, Old LJ, Schreiber RD. IFNgamma and lymphocytes prevent primary tumour development and shape tumour immunogenicity. Nature 2001; 410(6832): 1107–1111

[94] Lv H, Lv G, Chen C, Zong Q, Jiang G, Ye D, Cui X, He Y, Xiang W, Han Q, Tang L, Yang W, Wang H. NAD⁺ metabolism maintains inducible PD-L1 expression to drive tumor immune evasion. Cell Metab 2021; 33(1): 110–127.e5

[95] Dai X, Guo Y, Hu Y, Bao X, Zhu X, Fu Q, Zhang H, Tong Z, Liu L, Zheng Y, Zhao P, Fang W. Immunotherapy for targeting cancer stem cells in hepatocellular carcinoma. Theranostics 2021; 11(7): 3489–3501

[96] Hsu JM, Xia W, Hsu YH, Chan LC, Yu WH, Cha JH, Chen CT, Liao HW, Kuo CW, Khoo KH, Hsu JL, Li CW, Lim SO, Chang SS, Chen YC, Ren GX, Hung MC. STT3-dependent PD-L1 accumulation on cancer stem cells promotes immune evasion. Nat Commun 2018; 9(1): 1908

[97] Morrison BJ, Steel JC, Morris JC. Reduction of MHC-I expression limits T-lymphocyte-mediated killing of cancer-initiating cells. BMC Cancer 2018; 18(1): 469

[98] Dianat-Moghadam H, Rokni M, Marofi F, Panahi Y, Yousefi M. Natural killer cell-based immunotherapy: from transplantation toward targeting cancer stem cells. J Cell Physiol 2018; 234(1): 259–273

[99] Cheung PF, Yip CW, Wong NC, Fong DY, Ng LW, Wan AM, Wong CK, Cheung TT, Ng IO, Poon RT, Fan ST, Cheung ST. Granulin-epithelin precursor renders hepatocellular carcinoma cells resistant to natural killer cytotoxicity. Cancer Immunol Res 2014; 2(12): 1209–1219

[100] Lam KH, Ma S. Noncellular components in the liver cancer stem cell niche: biology and potential clinical implications. Hepatology 2023; 78(3): 991–1005

[101] You Y, Zheng Q, Dong Y, Xie X, Wang Y, Wu S, Zhang L, Wang Y, Xue T, Wang Z, Chen R, Wang Y, Cui J, Ren Z. Matrix stiffness-mediated effects on stemness characteristics occurring in HCC cells. Oncotarget 2016; 7(22): 32221–32231

[102] Kohga K, Tatsumi T, Takehara T, Tsunematsu H, Shimizu S, Yamamoto M, Sasakawa A, Miyagi T, Hayashi N. Expression of CD133 confers malignant potential by regulating metalloproteinases in human hepatocellular carcinoma. J Hepatol 2010; 52(6): 872–879

[103] Zhao W, Lv M, Yang X, Zhou J, Xing B, Zhang Z. Liver tumor-initiating cells initiate the formation of a stiff cancer stem cell microenvironment niche by secreting LOX. Carcinogenesis 2022; 43(8): 766–778

[104] Cui CP, Wong CC, Kai AK, Ho DW, Lau EY, Tsui YM, Chan LK, Cheung TT, Chok KS, Chan ACY, Lo RC, Lee JM, Lee TK, Ng IOL. SENP1 promotes hypoxia-induced cancer stemness by HIF-1α deSUMOylation and SENP1/HIF-1α positive feedback loop. Gut 2017; 66(12): 2149–2159

[105] Liu G, Luo Q, Li H, Liu Q, Ju Y, Song G. Increased oxidative phosphorylation is required for stemness maintenance in liver cancer stem cells from hepatocellular carcinoma cell line HCCLM3 cells. Int J Mol Sci 2020; 21(15): 5276

[106] Liu Y, Ren H, Zhou Y, Shang L, Zhang Y, Yang F, Shi X. The hypoxia conditioned mesenchymal stem cells promote hepatocellular carcinoma progression through YAP mediated lipogenesis reprogramming. J Exp Clin Cancer Res 2019; 38(1): 228

[107] Ling S, Shan Q, Zhan Q, Ye Q, Liu P, Xu S, He X, Ma J, Xiang J, Jiang G, Wen X, Feng Z, Wu Y, Feng T, Xu L, Chen K, Zhang X, Wei R, Zhang C, Cen B, Xie H, Song P, Liu J, Zheng S, Xu X. USP22 promotes hypoxia-induced hepatocellular carcinoma stemness by a HIF1α/USP22 positive feedback loop upon TP53 inactivation. Gut 2020; 69(7): 1322–1334

[108] Chisari A, Golán I, Campisano S, Gélabert C, Moustakas A, Sancho P, Caja L. Glucose and amino acid metabolic dependencies linked to stemness and metastasis in different aggressive cancer types. Front Pharmacol 2021; 12: 723798

[109] Lin SH, Liu T, Ming X, Tang Z, Fu L, Schmitt-Kopplin P, Kanawati B, Guan XY, Cai Z. Regulatory role of hexosamine biosynthetic pathway on hepatic cancer stem cell marker CD133 under low glucose conditions. Sci Rep 2016; 6(1): 21184

[110] Fan H, Zhang H, Pascuzzi PE, Andrisani O. Hepatitis B virus X protein induces EpCAM expression via active DNA demethylation directed by RelA in complex with EZH2 and TET2. Oncogene 2016; 35(6): 715–726

[111] Ali N, Allam H, May R, Sureban SM, Bronze MS, Bader T, Umar S, Anant S, Houchen CW. Hepatitis C virus-induced cancer stem cell-like signatures in cell culture and murine tumor xenografts. J Virol 2011; 85(23): 12292–12303

[112] Zhu M, Li W, Lu Y, Dong X, Lin B, Chen Y, Zhang X, Guo J, Li M. HBx drives alpha fetoprotein expression to promote initiation of liver cancer stem cells through activating PI3K/AKT signal pathway. Int J Cancer 2017; 140(6): 1346–1355

[113] Shrivastava S, Mukherjee A, Ray R, Ray RB. Hepatitis C virus induces interleukin-1β (IL-1β)/IL-18 in circulatory and resident liver macrophages. J Virol 2013; 87(22): 12284–12290

[114] Lin X, Zuo S, Luo R, Li Y, Yu G, Zou Y, Zhou Y, Liu Z, Liu Y, Hu Y, Xie Y, Fang W, Liu Z. HBX-induced miR-5188 impairs FOXO1 to stimulate β-catenin nuclear translocation and promotes tumor stemness in hepatocellular carcinoma. Theranostics 2019; 9(25): 7583–7598

[115] Uthaya Kumar DB, Chen CL, Liu JC, Feldman DE, Sher LS, French S, DiNorcia J, French SW, Naini BV, Junrungsee S, Agopian VG, Zarrinpar A, Machida K. TLR4 signaling via NANOG cooperates with STAT3 to activate twist1 and promote formation of tumor-initiating stem-like cells in livers of mice. Gastroenterology 2016; 150(3): 707–719

[116] Fan H, Cui Z, Zhang H, Mani SK, Diab A, Lefrancois L, Fares N, Merle P, Andrisani O. DNA demethylation induces SALL4 gene re-expression in subgroups of hepatocellular carcinoma associated with Hepatitis B or C virus infection. Oncogene 2017; 36(17): 2435–2445

[117] Broz ML, Krummel MF. The emerging understanding of myeloid cells as partners and targets in tumor rejection. Cancer Immunol Res 2015; 3(4): 313–319

[118] Gabrilovich DI, Ostrand-Rosenberg S, Bronte V. Coordinated regulation of myeloid cells by tumours. Nat Rev Immunol 2012; 12(4): 253–268

[119] Engblom C, Pfirschke C, Pittet MJ. The role of myeloid cells in cancer therapies. Nat Rev Cancer 2016; 16(7): 447–462

[120] Gentek R, Molawi K, Sieweke MH. Tissue macrophage identity and self-renewal. Immunol Rev 2014; 262(1): 56–73

[121] Wu K, Lin K, Li X, Yuan X, Xu P, Ni P, Xu D. Redefining tumor-associated macrophage subpopulations and functions in the tumor microenvironment. Front Immunol 2020; 11: 1731

[122] Jhunjhunwala S, Hammer C, Delamarre L. Antigen presentation in cancer: insights into tumour immunogenicity and immune evasion. Nat Rev Cancer 2021; 21(5): 298–312

[123] Xiao P, Long X, Zhang L, Ye Y, Guo J, Liu P, Zhang R, Ning J, Yu W, Wei F, Yu J. Neurotensin/IL-8 pathway orchestrates local inflammatory response and tumor invasion by inducing M2 polarization of tumor-associated macrophages and epithelial-mesenchymal transition of hepatocellular carcinoma cells. OncoImmunology 2018; 7(7): e1440166

[124] Zhu F, Li X, Chen S, Zeng Q, Zhao Y, Luo F. Tumor-associated macrophage or chemokine ligand CCL17 positively regulates the tumorigenesis of hepatocellular carcinoma. Med Oncol 2016; 33(2): 17

[125] Yan W, Liu X, Ma H, Zhang H, Song X, Gao L, Liang X, Ma C. Tim-3 fosters HCC development by enhancing TGF-β-mediated alternative activation of macrophages. Gut 2015; 64(10): 1593–1604

[126] Fu XT, Dai Z, Song K, Zhang ZJ, Zhou ZJ, Zhou SL, Zhao YM, Xiao YS, Sun QM, Ding ZB, Fan J. Macrophage-secreted IL-8 induces epithelial-mesenchymal transition in hepatocellular carcinoma cells by activating the JAK2/STAT3/Snail pathway. Int J Oncol 2015; 46(2): 587–596

[127] Peng ZP, Jiang ZZ, Guo HF, Zhou MM, Huang YF, Ning WR, Huang JH, Zheng L, Wu Y. Glycolytic activation of monocytes regulates the accumulation and function of neutrophils in human hepatocellular carcinoma. J Hepatol 2020; 73(4): 906–917

[128] Wang Y, Wang B, Xiao S, Li Y, Chen Q. miR-125a/b inhibits tumor-associated macrophages mediated in cancer stem cells of hepatocellular carcinoma by targeting CD90. J Cell Biochem 2019; 120(3): 3046–3055

[129] Iliopoulos D, Hirsch HA, Struhl K. An epigenetic switch involving NF-κB, Lin28, Let-7 MicroRNA, and IL6 links inflammation to cell transformation. Cell 2009; 139(4): 693–706

[130] Barkal AA, Brewer RE, Markovic M, Kowarsky M, Barkal SA, Zaro BW, Krishnan V, Hatakeyama J, Dorigo O, Barkal LJ, Weissman IL. CD24 signalling through macrophage Siglec-10 is a target for cancer immunotherapy. Nature 2019; 572(7769): 392–396

[131] Arvanitakis K, Mitroulis I, Germanidis G. Tumor-associated neutrophils in hepatocellular carcinoma pathogenesis, prognosis, and therapy. Cancers (Basel) 2021; 13(12): 2899

[132] Song M, He J, Pan QZ, Yang J, Zhao J, Zhang YJ, Huang Y, Tang Y, Wang Q, He J, Gu J, Li Y, Chen S, Zeng J, Zhou ZQ, Yang C, Han Y, Chen H, Xiang T, Weng DS, Xia JC. Cancer-associated fibroblast-mediated cellular crosstalk supports hepatocellular carcinoma progression. Hepatology 2021; 73(5): 1717–1735

[133] Ma T, Renz BW, Ilmer M, Koch D, Yang Y, Werner J, Bazhin AV. Myeloid-derived suppressor cells in solid tumors. Cells 2022; 11(2): 310

[134] Tcyganov E, Mastio J, Chen E, Gabrilovich DI. Plasticity of myeloid-derived suppressor cells in cancer. Curr Opin Immunol 2018; 51: 76–82

[135] Ostrand-Rosenberg S. Myeloid-derived suppressor cells: more mechanisms for inhibiting antitumor immunity. Cancer Immunol Immunother 2010; 59(10): 1593–1600

[136] Xu M, Zhao Z, Song J, Lan X, Lu S, Chen M, Wang Z, Chen W, Fan X, Wu F, Chen L, Tu J, Ji J. Interactions between interleukin-6 and myeloid-derived suppressor cells drive the chemoresistant phenotype of hepatocellular cancer. Exp Cell Res 2017; 351(2): 142–149

[137] Lee WC, Hsu PY, Hsu HY. Stem cell factor produced by tumor cells expands myeloid-derived suppressor cells in mice. Sci Rep 2020; 10(1): 11257

[138] Liu M, Zhou J, Liu X, Feng Y, Yang W, Wu F, Cheung OK, Sun H, Zeng X, Tang W, Mok MTS, Wong J, Yeung PC, Lai PBS, Chen Z, Jin H, Chen J, Chan SL, Chan AWH, To KF, Sung JJY, Chen M, Cheng AS. Targeting monocyte-intrinsic enhancer reprogramming improves immunotherapy efficacy in hepatocellular carcinoma. Gut 2020; 69(2): 365–379

[139] He Q, Liu M, Huang W, Chen X, Zhang B, Zhang T, Wang Y, Liu D, Xie M, Ji X, Sun M, Tian D, Xia L. IL-1β-induced elevation of solute carrier family 7 member 11 promotes hepatocellular carcinoma metastasis through up-regulating programmed death ligand 1 and colony-stimulating factor 1. Hepatology 2021; 74(6): 3174–3193

[140] Lin Y, Cai Q, Chen Y, Shi T, Liu W, Mao L, Deng B, Ying Z, Gao Y, Luo H, Yang X, Huang X, Shi Y, He R. CAFs shape myeloid-derived suppressor cells to promote stemness of intrahepatic cholangiocarcinoma through 5-lipoxygenase. Hepatology 2022; 75(1): 28–42

[141] Wang S, Sun J, Chen K, Ma P, Lei Q, Xing S, Cao Z, Sun S, Yu Z, Liu Y, Li N. Perspectives of tumor-infiltrating lymphocyte treatment in solid tumors. BMC Med 2021; 19(1): 140

[142] Idos GE, Kwok J, Bonthala N, Kysh L, Gruber SB, Qu C. The prognostic implications of tumor infiltrating lymphocytes in colorectal cancer: a systematic review and meta-analysis. Sci Rep 2020; 10(1): 3360

[143] Ren L, Yu Y, Wang L, Zhu Z, Lu R, Yao Z. Hypoxia-induced CCL28 promotes recruitment of regulatory T cells and tumor growth in liver cancer. Oncotarget 2016; 7(46): 75763–75773

[144] Yang P, Li QJ, Feng Y, Zhang Y, Markowitz GJ, Ning S, Deng Y, Zhao J, Jiang S, Yuan Y, Wang HY, Cheng SQ, Xie D, Wang XF. TGF-β-miR-34a-CCL22 signaling-induced Treg cell recruitment promotes venous metastases of HBV-positive hepatocellular carcinoma. Cancer Cell 2012; 22(3): 291–303

[145] Kurebayashi Y, Ojima H, Tsujikawa H, Kubota N, Maehara J, Abe Y, Kitago M, Shinoda M, Kitagawa Y, Sakamoto M. Landscape of immune microenvironment in hepatocellular carcinoma and its additional impact on histological and molecular classification. Hepatology 2018; 68(3): 1025–1041

[146] Calderaro J, Rousseau B, Amaddeo G, Mercey M, Charpy C, Costentin C, Luciani A, Zafrani ES, Laurent A, Azoulay D, Lafdil F, Pawlotsky JM. Programmed death ligand 1 expression in hepatocellular carcinoma: relationship with clinical and pathological features. Hepatology 2016; 64(6): 2038–2046

[147] Hsu JM, Xia W, Hsu YH, Chan LC, Yu WH, Cha JH, Chen CT, Liao HW, Kuo CW, Khoo KH, Hsu JL, Li CW, Lim SO, Chang SS, Chen YC, Ren GX, Hung MC. STT3-dependent PD-L1 accumulation on cancer stem cells promotes immune evasion. Nat Commun 2018; 9(1): 1908

[148] Chan LC, Li CW, Xia W, Hsu JM, Lee HH, Cha JH, Wang HL, Yang WH, Yen EY, Chang WC, Zha Z, Lim SO, Lai YJ, Liu C, Liu J, Dong Q, Yang Y, Sun L, Wei Y, Nie L, Hsu JL, Li H, Ye Q, Hassan MM, Amin HM, Kaseb AO, Lin X, Wang SC, Hung MC. IL-6/JAK1 pathway drives PD-L1 Y112 phosphorylation to promote cancer immune evasion. J Clin Invest 2019; 129(8): 3324–3338

[149] Wei Z, Jia J, Heng G, Xu H, Shan J, Wang G, Liu C, Xia J, Zhou H, Wu M, Yang Z, Wang M, Xiong Z, Huang H, Liu L, Qian C. Sirtuin-1/mitochondrial ribosomal protein S5 axis enhances the metabolic flexibility of liver cancer stem cells. Hepatology 2019; 70(4): 1197–1213

[150] Ruiz de Galarreta M, Bresnahan E, Molina-Sánchez P, Lindblad KE, Maier B, Sia D, Puigvehi M, Miguela V, Casanova-Acebes M, Dhainaut M, Villacorta-Martin C, Singhi AD, Moghe A, von Felden J, Tal Grinspan L, Wang S, Kamphorst AO, Monga SP, Brown BD, Villanueva A, Llovet JM, Merad M, Lujambio A. β-catenin activation promotes immune escape and resistance to anti-PD-1 therapy in hepatocellular carcinoma. Cancer Discov 2019; 9(8): 1124–1141

[151] Zhu GQ, Wang Y, Wang B, Liu WR, Dong SS, Chen EB, Cai JL, Wan JL, Du JX, Song LN, Chen SP, Yu L, Zhou ZJ, Wang Z, Zhou J, Shi YH, Fan J, Dai Z. Targeting HNRNPM inhibits cancer stemness and enhances antitumor immunity in Wnt-activated hepatocellular carcinoma. Cell Mol Gastroenterol Hepatol 2022; 13(5): 1413–1447

[152] Rodríguez MM, Fiore E, Bayo J, Atorrasagasti C, García M, Onorato A, Domínguez L, Malvicini M, Mazzolini G. 4Mu decreases CD47 expression on hepatic cancer stem cells and primes a potent antitumor T cell response induced by interleukin-12. Mol Ther 2018; 26(12): 2738–2750

[153] Cheung ST, Cheung PFY, Cheng CKC, Wong NCL, Fan ST. Granulin-epithelin precursor and ATP-dependent binding cassette (ABC)B5 regulate liver cancer cell chemoresistance. Gastroenterology 2011; 140(1): 344–355

[154] Cheung PF, Yip CW, Wong NC, Fong DY, Ng LW, Wan AM, Wong CK, Cheung TT, Ng IO, Poon RT, Fan ST, Cheung ST. Granulin-epithelin precursor renders hepatocellular carcinoma cells resistant to natural killer cytotoxicity. Cancer Immunol Res 2014; 2(12): 1209–1219

[155] Park DJ, Sung PS, Kim JH, Lee GW, Jang JW, Jung ES, Bae SH, Choi JY, Yoon SK. EpCAM-high liver cancer stem cells resist natural killer cell-mediated cytotoxicity by upregulating CEACAM1. J Immunother Cancer 2020; 8(1): e000301

[156] Zhang PF, Gao C, Huang XY, Lu JC, Guo XJ, Shi GM, Cai JB, Ke AW. Cancer cell-derived exosomal circUHRF1 induces natural killer cell exhaustion and may cause resistance to anti-PD1 therapy in hepatocellular carcinoma. Mol Cancer 2020; 19(1): 110

[157] Ma Y, Shurin GV, Peiyuan Z, Shurin MR. Dendritic cells in the cancer microenvironment. J Cancer 2013; 4(1): 36–44

[158] Ng KY, Chai S, Tong M, Guan XY, Lin CH, Ching YP, Xie D, Cheng AS, Ma S. C-terminal truncated hepatitis B virus X protein promotes hepatocellular carcinogenesis through induction of cancer and stem cell-like properties. Oncotarget 2016; 7(17): 24005–24017

[159] Weiner GJ. Building better monoclonal antibody-based therapeutics. Nat Rev Cancer 2015; 15(6): 361–370

[160] Sun YF, Xu Y, Yang XR, Guo W, Zhang X, Qiu SJ, Shi RY, Hu B, Zhou J, Fan J. Circulating stem cell-like epithelial cell adhesion molecule-positive tumor cells indicate poor prognosis of hepatocellular carcinoma after curative resection. Hepatology 2013; 57(4): 1458–1468

[161] Zhang P, Shi B, Gao H, Jiang H, Kong J, Yan J, Pan X, Li K, Zhang P, Yao M, Yang S, Gu J, Wang H, Li Z. An EpCAM/CD3 bispecific antibody efficiently eliminates hepatocellular carcinoma cells with limited galectin-1 expression. Cancer Immunol Immunother 2014; 63(2): 121–132

[162] Huang J, Li C, Wang Y, Lv H, Guo Y, Dai H, Wicha MS, Chang AE, Li Q. Cytokine-induced killer (CIK) cells bound with anti-CD3/anti-CD133 bispecific antibodies target CD133(high) cancer stem cells in vitro and in vivo. Clin Immunol 2013; 149(1): 156–168

[163] Xiao Z, Chung H, Banan B, Manning PT, Ott KC, Lin S, Capoccia BJ, Subramanian V, Hiebsch RR, Upadhya GA, Mohanakumar T, Frazier WA, Lin Y, Chapman WC. Antibody mediated therapy targeting CD47 inhibits tumor progression of hepatocellular carcinoma. Cancer Lett 2015; 360(2): 302–309

[164] Wang L, Su W, Liu Z, Zhou M, Chen S, Chen Y, Lu D, Liu Y, Fan Y, Zheng Y, Han Z, Kong D, Wu JC, Xiang R, Li Z. CD44 antibody-targeted liposomal nanoparticles for molecular imaging and therapy of hepatocellular carcinoma. Biomaterials 2012; 33(20): 5107–5114

[165] Sun F, Wang T, Jiang J, Wang Y, Ma Z, Li Z, Han Y, Pan M, Cai J, Wang M, Zhang J. Engineering a high-affinity humanized anti-CD24 antibody to target hepatocellular carcinoma by a novel CDR grafting design. Oncotarget 2017; 8(31): 51238–51252

[166] Hirohashi Y, Torigoe T, Inoda S, Takahashi A, Morita R, Nishizawa S, Tamura Y, Suzuki H, Toyota M, Sato N. Immune response against tumor antigens expressed on human cancer stem-like cells/tumor-initiating cells. Immunotherapy 2010; 2(2): 201–211

[167] Saijo H, Hirohashi Y, Torigoe T, Kochin V, Takahashi H, Sato N. Cytotoxic T lymphocytes: the future of cancer stem cell eradication?. Immunotherapy 2013; 5(6): 549–551

[168] Deng Z, Wu Y, Ma W, Zhang S, Zhang YQ. Adoptive T-cell therapy of prostate cancer targeting the cancer stem cell antigen EpCAM. BMC Immunol 2015; 16(1): 1

[169] Alhabbab RY. Targeting cancer stem cells by genetically engineered chimeric antigen receptor T cells. Front Genet 2020; 11: 312

[170] Wang Y, Chen M, Wu Z, Tong C, Dai H, Guo Y, Liu Y, Huang J, Lv H, Luo C, Feng KC, Yang QM, Li XL, Han W. CD133-directed CAR T cells for advanced metastasis malignancies: a phase I trial. OncoImmunology 2018; 7(7): e1440169

[171] Dai H, Tong C, Shi D, Chen M, Guo Y, Chen D, Han X, Wang H, Wang Y, Shen P. Efficacy and biomarker analysis of CD133-directed CAR T cells in advanced hepatocellular carcinoma: a single-arm, open-label, phase II trial. OncoImmunology 2020; 9(1): 1846926

[172] Zhai Y, He K, Huang L, Shang X, Wang G, Yuan G, Han ZG. DLK1-directed chimeric antigen receptor T-cell therapy for hepatocellular carcinoma. Liver Int 2022; 42(11): 2524–2537

[173] Dal Bo M, De Mattia E, Baboci L, Mezzalira S, Cecchin E, Assaraf YG, Toffoli G. New insights into the pharmacological, immunological, and CAR-T-cell approaches in the treatment of hepatocellular carcinoma. Drug Resist Updat 2020; 51: 100702

[174] Morgan RA, Yang JC, Kitano M, Dudley ME, Laurencot CM, Rosenberg SA. Case report of a serious adverse event following the administration of T cells transduced with a chimeric antigen receptor recognizing ERBB2. Mol Ther 2010; 18(4): 843–851

[175] Sun S, Hao H, Yang G, Zhang Y, Fu Y. Immunotherapy with CAR-modified T cells: toxicities and overcoming strategies. J Immunol Res 2018; 2018: 2386187

[176] Choi YJ, Park SJ, Park YS, Park HS, Yang KM, Heo K. EpCAM peptide-primed dendritic cell vaccination confers significant anti-tumor immunity in hepatocellular carcinoma cells. PLoS One 2018; 13(1): e0190638

[177] Sun JC, Pan K, Chen MS, Wang QJ, Wang H, Ma HQ, Li YQ, Liang XT, Li JJ, Zhao JJ, Chen YB, Pang XH, Liu WL, Cao Y, Guan XY, Lian QZ, Xia JC. Dendritic cells-mediated CTLs targeting hepatocellular carcinoma stem cells. Cancer Biol Ther 2010; 10(4): 368–375

[178] Pan QZ, Pan K, Wang QJ, Weng DS, Zhao JJ, Zheng HX, Zhang XF, Jiang SS, Lv L, Tang Y, Li YQ, He J, Liu Q, Chen CL, Zhang HX, Xia JC. Annexin A3 as a potential target for immunotherapy of liver cancer stem-like cells. Stem Cells 2015; 33(2): 354–366

[179] Wang Y, Zhao Q, Zhao B, Zheng Y, Zhuang Q, Liao N, Wang P, Cai Z, Zhang D, Zeng Y, Liu X. Remodeling tumor-associated neutrophils to enhance dendritic cell-based HCC neoantigen nano-vaccine efficiency. Adv Sci (Weinh) 2022; 9(11): e2105631

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