Macrophages are a population of immune cells with a high degree of phenotypic and functional heterogeneity.1 Besides their roles in homeostasis, tissue repair, and development, roles of macrophages in tumor and leukemia become the focus.2 In solid tumor tissues, there are various immune cells and stroma cells in addition to tumor cells, such as macrophages, neutrophils, mast cells, fat cells, and fibroblasts.3 Tumor-associated macrophages (TAMs) promote tumor progression, and their accumulation in the tumors correlates with poor prognosis in many cancer types.4 In hematological malignancies, such as lymphoma, myeloma, and leukemia, macrophages infiltrate into disease microenvironment, obtain specific activation phenotypes, and participate in disease progression.5 Compared with solid tumors, leukemia has different clinical features. During leukemogenesis, leukemia cells outcompete their normal counterparts (hematopoietic stem cells [HSCs]) in bone marrow (BM), which is the major site for hematopoiesis.6–8 Macrophages, as an important component in BM microenvironment, have remarkable plasticity. Their functional phenotype is controlled by microenvironmental signals. Macrophages in leukemia microenvironment are called leukemia-associated macrophages (LAMs). They are involved in disease progression.
2 HETEROGENEITY OF MACROPHAGES
Under physiological conditions, macrophages are heterogeneous in several aspects, that is, tissue-specific phenotypes/functions, developmental origins, and polarization by different stimuli.9 It is well-established that tissue-specific microenvironments give rise to the heterogeneity of tissue macrophages. The phenotypic markers and transcription regulators of tissue-resident macrophages in different macroenvironments are summarized in Table 1.
Recent evidence reveals that macrophages in adult have different developmental origins. In general, adult tissue-resident macrophages have three origins, that is, yolk sac, fetal liver, and circulating monocytes. The origin of tissue macrophages varies among tissues. Tissue-resident macrophages in brain, known as microglia, may arise only from yolk sac macrophages. Langerhans, alveolar macrophages, and Kupffer cells originate from both yolk sac and fetal liver monocytes. In heart-, pancreas-, and gut tissue-resident macrophages of fetal origin will gradually be replaced by BM- and monocyte-derived macrophages.10 Although they have almost identical phenotype and function in the same microenvironment, macrophages from different origins may have different developmental imprints. For example, Siglec-F and CCR2 can be used to identify alveolar macrophages with embryonic and monocytic origins since embryonic origin macrophages are Siglec-Fhi and monocytic origin ones are CCR2hi.10
Macrophages have remarkable plasticity that allows them to respond efficiently to numerous environmental signals. They are heterogeneous in activation phenotype. The classically activated macrophages (M1) and alternatively activated macrophages (M2) are well-accepted phenotypes of macrophage activation. M1 macrophages are polarized by LPS and IFN-γ. They express high level of iNOS and secrete proinflammatory cytokines and express inflammatory chemokines such as CXCL9 and CXCL10. M2 macrophages are polarized by IL-4 and IL-6. They express high level of Arg1 and secrete TGF-β, IL-10, arginase, and metalloprotease to participate in immunosuppression.11 However, macrophage phenotypes may be more appropriately described as a continuum of functional states. M1 and M2 macrophages represent only the two extremes of macrophage activation phenotypes.12 Nowadays, M2 macrophages refer to all non-M1 macrophages and are further divided into M2a to M2d subtypes. TAMs are regarded as an M2-like phenotype, M2d.13 M1- and M2-associated genes have been suggested. Though no gene expression is unique for either M1 or M2 macrophages, high-level expression of some genes is commonly observed in M1 or M2 macrophages. For example, iNOS and Arg1 are frequently used as markers for M1 and M2 macrophages, respectively. In general, M1 macrophages have antitumor effects, whereas M2 macrophages have protumor effects. A color wheel hypothesis has also been proposed based on three fundamental populations of macrophages that include classically activated macrophages, wound-healing macrophages, and regulatory macrophages.14
Under pathological conditions, such as tumor and leukemia microenvironments, macrophages retain their plasticity and respond to disease microenvironmental signals.1 Macrophages recruited into tumor microenvironment are educated to an M2-like phenotype.15 TAMs, as an important component of the tumor microenvironment, involve in the tumor-related immunosuppression.16 M2-like TAMs promote tumor growth, invasion, and metastasis by secreting growth factors, cytokines such as angiogenesis factors, and various proteases.17 LAMs express both M1- and M2-associated genes. Similar to TAMs, LAMs consist of M1- and M2-like subpopulations based on CD206 expression.18 Macrophages in chronic lymphocytic leukemia (CLL) are known as “nanny-like cells” (NLCs), which exhibit similar phenotype with TAMs in B-cell lymphoma.19
3 CHARACTERISTICS OF LAMS IN LEUKEMIA MICROENVIRONMENT
In hematological malignancies, TAMs in lymphoma and myeloma are widely studied and in many cases TAM counts in tissue sections correlate with poor prognosis.20,21 In recent years, studies on phenotype and function of macrophages in leukemia microenvironment reveal their characteristics.
From different leukemia models, increase of LAMs in hematopoietic microenvironment is observed during the development of leukemia, especially early and middle stages of leukemia. In nonirradiated Notch1-induced mouse T-cell acute lymphoblastic leukemia (T-ALL) and MLL-AF9-induced mouse acute myeloid leukemia (AML) models, macrophages infiltrated into BM and spleen increase during early stage of leukemia progression and decrease thereafter.18,22 In another report, in which macrophages in AML are also defined as AML-associated macrophages (AAMs), increase of macrophages in BM is detected in AML patients. This phenomenon is also observed in NUP98-HOXD13 mouse AML model.23 These observations suggest that noninfectious inflammation occur in the development of leukemia and macrophages play pathological roles in the progression of leukemia.
Although LAMs are generally considered as M2-like macrophages, the activation phenotype of LAMs is heterogeneous. In Notch1-induced T-ALL and MLL-AF9-induced AML, LAMs from different organs express both M1 and M2 macrophage activation-associated molecules.18,22 AAMs in leukemia burden mice express higher levels of Arg1 and lower levels of IL-6 and Nos2, exhibiting M2 like phenotype.23 LAMs in T-ALL model consist of CD206+ and CD206− subpopulations, especially at early stage of leukemia. Two subpopulations express both M1- and M2-associated genes. But CD206+ LAMs expressed higher levels of both M1- and M2-associated genes than CD206− LAMs.18 Similar results are also observed in AML model, and gene expression profiles of LAMs from BM and spleen in T-ALL and AML mice show that they have distinct gene expression signature.22 During the development of CLL, leukemia cells secrete chemokines to induce peripheral monocytes migration to them. And infiltrative monocytes differentiate into NLCs with M2 phenotype, which express high level of CD14, CD68, CD163, and CD206.24
In physiological state, macrophages showed tissue-specific gene expression profiles determined by gene-enhancer landscapes shaped by microenvironments.25 LAMs present organ-specific activation as well. Peritoneal tissue-derived LAMs simultaneously expressed high-level iNOS and Arg1, which was not commonly observed in macrophages from other tissues and origins. A study comparing the activation phenotype of LAMs from BM, spleen, and peritoneal tissue in T-ALL model demonstrates that considerable phenotypic diversities are detected among those LAMs, although they all express lower level of CSF-1, TGF-β1, and VEGFα than TAMs.26 A study comparing the activation phenotype of LAMs from BM and spleen in T-ALL and AML models reveals that LAMs in spleen have more M2 characteristics, whereas LAMs in BM have more M1 characteristics. IRF7 promoted M1 characteristics through activation of SAPK/JNK pathway in macrophages.22 LAMs in liver exhibited a more M1-like phenotype distinct from LAMs in BM or spleen. They express higher level of CCL5, iNOS, IL12β, TNFα, MCSF, MMP9, TGFβ, and VEGFα, while lower level of IL1-β, CD206, and IL 10 than that in BM and spleen LAMs.27 Furthermore, LAM subpopulations in different organs show considerable diversity. For instance, the expression of IL-1β, iNOS, IL-6, Arg1, and IL-10 from CD206+ LAMs in spleen is higher than that in BM.18 The phenotypic characteristics of macrophages in different types of leukemia are summarized in Table 2.
4 ROLES OF MACROPHAGES IN LEUKEMIA MICROENVIRONMENT
It is well known that TAMs infiltrate into tumors and promote tumor progression through affecting proliferation of tumor cells, angiogenesis, immunosuppression, and so on. Accumulation of TAMs is correlated with worse prognosis in many cancers, including lymphoma and myeloma.20,28–31 Similar to TAMs, LAMs are an adverse factor in leukemia development. Patients with higher level of CD163 expression have a shorter survival than those with lower level of CD163.22 In T-ALL and AML mouse models, LAMs promote the proliferation of leukemia cells.18,22 The leukemia microenvironment polarizes AAMs to M2 phenotype through regulating Gfi1, which supports the growth of AML cells.23 CD14+ blood cells from healthy donors differentiate into NLCs when cocultured with CLL B cells and protect CLL cells from apoptosis.24 CLL cells secrete IL-4,31 IL-13,32 IL-10 33, and other cytokines and promote NLCs/TAMs polarization to M2 phenotype, while M2-like NLCs/TAMs inhibits T cell infiltration and mediates local immunosuppression. NLCs/TAMs also promote the survival of CLL cells by secreting IGF1, IL-8, CCL2, and CXCL12.34 In the Eu-TCL1 transgenic CLL model, macrophages support the growth of CLL cells in mice,35 protumorigenic and immunosuppressive properties of macrophages can be modulated through the PI3K/mTOR signaling pathway.36
5 LAMS AND TAMS
In leukemia and tumor microenvironments, proliferation of malignant cells largely depends on local malignant niches. LAMs and TAMs in respective niche exhibit similar but different phenotypic and functional characteristics. Several similarities are found between LAMs and TAMs. First, increase of LAMs and TAMs are commonly detected in malignant microenvironments when compared with their normal counterparts.18,23,37,38 Second, LAM and TAM populations are heterogeneous and can be subdivided into different subpopulations. TAMs in tumor microenvironment consist of M1-like TAMs and M2-like TAMs. The phenotype of TAMs varies in different areas of tumor tissues. In a breast cancer model, TAMs localized in normoxic sites express M1 markers and antiangiogenic chemokines, whereas TAMs in hypoxic sites express M2 markers and have proangiogenic effects.39 M2 like TAMs have the effects of immunosuppression, angiogenesis, and promoting the metastasis.40–42 In leukemia microenvironment, LAMs consist of CD206+ and CD206− subpopulations. Both CD206+ and CD206− LAMs have proleukemia effects.18 Third, LAMs and TAMs have tissue-specific phenotypes. TAMs in metastatic tumor exhibit more M2 phenotype than primary lesion.43 As already discussed, LAMs from BM, spleen, liver, and abdominal cavity have different phenotypic characteristics.18,22,26,27
The functional characteristics of LAMs and TAMs also have differences. Although both LAMs and TAMs promote malignant progression, the main mechanisms may be different. As angiogenesis is much more important in solid tumors, high-level expression of pro-angiogenetic factors, such as VEGFα, are frequently detected in TAMs, whereas it is not detected in LAMs in several tissues from different leukemia models.18,22,26,27 TAMs also promote tumor progression through immunosuppression and promoting invasion and metastasis of tumor cells. In CLL microenvironment, M2-like NLCs/TAMs play important roles in immunosuppression through secreting inflammatory suppressing cytokines, such as IL-4, IL-10, and IL-13.31–33 It is still unclear whether immunosuppression effects play important roles in other types of leukemia. Furthermore, the role of LAMs in invasion and metastasis of leukemia cells has not been established. In the CLL and AML model, LAMs promote proliferation or inhibit apoptosis of leukemia cells directly in AML microenvironment.18,23,34,35
6 TARGETING MACROPHAGE FOR LEUKEMIA THERAPY AND FUTURE EXPECTATION
Macrophage infiltration has been shown to be an independent poor prognostic factor in several cancer types as well as in leukemia.44 Hence, more attention has been paid to develop immunotherapies targeting TAMs and LAMs. TAMs-targeted therapies have been extensively studied in solid tumors. Intervention strategies include depleting TAMs, blocking their protumor signaling, and restoring their immune-stimulatory properties.41 With deep understanding of macrophage in hematological malignancies, strategies for targeting macrophages have been suggested and explored for therapy applications. In CLL mouse model, depletion of CLL-associated patrolling monocytes and macrophages using liposomal clodronate results in delay of disease development and repairs immune dysfunction.45 Targeting macrophages by either CSF1R signaling blockade or liposomal clodronate-mediated depletion has marked inhibitory effects on established leukemia.35 Interaction between SIRPα on macrophages and CD47 on AML cells is critical for leukemic engraftment and evasion of immune surveillance. Disruption of SIRPα-CD47 interaction by SIRPα-Fc fusion protein impairs AML engraftment and does not adversely affect normal hematopoiesis.46 Similar to CAR-T cell therapies, modified macrophages have been reported by introducing engineered chimeric antigen receptors for phagocytosis (CAR-Ps): Megf10 in macrophages. CAR-Ps result in specific engulfment of antigen-coated particles and human cancer cells.47 Macrophages-targeted therapies are promising strategies against leukemia and they may be practicable therapy options in clinical applications.
This work was supported by grants 81570153 and 81770183 from the National Natural Science Foundation of China (NSFC); programs 2016-I2M-2–006 and 2017-I2M-1–015 from the CAMS Innovation Fund for Medical Sciences (CIFMS); grant 17JCZDJC35000 from the Tianjin Natural Science Foundation. Z.G. is a recipient of the New Century Excellent Talents in University (NCET-08–0329).
. Stout RD, Jiang C, Matta B, Tietzel I, Watkins SK, Suttles J. Macrophages sequentially change their functional phenotype in response to changes in microenvironmental influences. J Immunol 2005;175(1):342.
. Wynn TA, Chawla A, Pollard JW. Macrophage biology in development, homeostasis and disease. Nature 2013;496(7446):445.
. Whiteside TL. The tumor microenvironment and its role in promoting tumor growth. Oncogene 2008;27(45):5904.
. Noy R, Pollard JW. Tumor-associated macrophages: from mechanisms to therapy. Immunity 2014;41(1):49.
. Komohara Y, Niino D, Ohnishi K, Ohshima K, Takeya M. Role of tumor-associated macrophages in hematological malignancies. Pathol Int 2015;65(4):170.
. Colmone A, Amorim M, Pontier AL, Wang S, Jablonski E, Sipkins DA. Leukemic cells create bone marrow niches that disrupt the behavior of normal hematopoietic progenitor cells. Science 2008;322(5909):1861.
. Hu X, Shen H, Tian C, et al. Kinetics of normal hematopoietic stem and progenitor cells in a Notch1-induced leukemia model. Blood 2009;114(18):3783.
. Cheng H, Hao S, Liu Y, et al. Leukemic marrow infiltration reveals a novel role for Egr3 as a potent inhibitor of normal hematopoietic stem cell proliferation. Blood 2015;126(11):1302.
. Varol C, Mildner A, Jung S. Macrophages: development and tissue specialization. Annu Rev Immunol 2015;33:643.
. Yao Y, Jeyanathan M, Haddadi S, et al. Induction of autonomous memory alveolar macrophages requires T cell help and is critical to trained immunity. Cell 2018;175(6):1634.
. Murray PJ, Allen JE, Biswas SK, et al. Macrophage activation and polarization: nomenclature and experimental guidelines. Immunity 2014;41(1):14.
. Gordon S. Alternative activation of macrophages. Nat Rev Immunol 2003;3(1):23.
. Martinez FO, Sica A, Mantovani A, Locati M. Macrophage activation and polarization. Front Biosci 2008;13:453.
. Mosser DM, Edwards JP. Exploring the full spectrum of macrophage activation. Nat Rev Immunol 2008;8(12):958.
. Mantovani A, Sozzani S, Locati M, Allavena P, Sica A. Macrophage polarization: tumor-associated macrophages as a paradigm for polarized M2 mononuclear phagocytes. Trends Immunol 2002;23(11):549.
. Liu Y, Cao X. The origin and function of tumor-associated macrophages. Cell Mol Immunol 2015;12(1):1.
. Qian BZ, Pollard JW. Macrophage diversity enhances tumor progression and metastasis. Cell 2010;141(1):39.
. Chen SY, Yang X, Feng WL, et al. Organ-specific microenvironment modifies diverse functional and phenotypic characteristics of leukemia-associated macrophages in mouse T cell acute lymphoblastic leukemia. J Immunol 2015;194(6):2919.
. Filip AA, Cisel B, Koczkodaj D, Wasik-Szczepanek E, Piersiak T, Dmoszynska A. Circulating microenvironment of CLL: are nurse-like cells related to tumor-associated macrophages? Blood Cells Mol Dis 2013;50(4):263.
. Steidl C, Lee T, Shah SP, et al. Tumor-associated macrophages and survival in classic Hodgkin's lymphoma. N Engl J Med 2010;362(10):875.
. Suyani E, Sucak GT, Akyurek N, et al. Tumor-associated macrophages as a prognostic parameter in multiple myeloma. Ann Hematol 2013;92(5):669.
. Yang X, Feng W, Wang R, et al. Repolarizing heterogeneous leukemia-associated macrophages with more M1 characteristics eliminates their pro-leukemic effects. Oncoimmunology 2018;7(4):e1412910.
. Al-Matary YS, Botezatu L, Opalka B, et al. Acute myeloid leukemia cells polarize macrophages towards a leukemia supporting state in a growth factor independence 1 dependent manner. Haematologica 2016;101(10):1216.
. Tsukada N, Burger JA, Zvaifler NJ, Kipps TJ. Distinctive features of “nurselike” cells that differentiate in the context of chronic lymphocytic leukemia. Blood 2002;99(3):1030.
. Gosselin D, Link VM, Romanoski CE, et al. Environment drives selection and function of enhancers controlling tissue-specific macrophage identities. Cell 2014;159(6):1327.
. Chen S, Yang X, Feng W, et al. Characterization of peritoneal leukemia-associated macrophages in Notch1-induced mouse T cell acute lymphoblastic leukemia. Mol Immunol 2017;81:35.
. Yang X, Feng W, Wang R, et al. Hepatic leukemia-associated macrophages exhibit a pro-inflammatory phenotype in Notch1-induced acute T cell leukemia. Immunobiology 2018;223(1):73.
. Troiano G, Caponio VCA, Adipietro I, et al. Prognostic significance of CD68(+) and CD163(+) tumor associated macrophages in head and neck squamous cell carcinoma: a systematic review and meta-analysis. Oral Oncol 2019;93:66.
. Qiu SQ, Waaijer SJH, Zwager MC, de Vries EGE, van der Vegt B, Schroder CP. Tumor-associated macrophages in breast cancer: innocent bystander or important player? Cancer Treat Rev 2018;70:178.
. Tan KL, Scott DW, Hong F, et al. Tumor-associated macrophages predict inferior outcomes in classic Hodgkin lymphoma: a correlative study from the E2496 Intergroup trial. Blood 2012;120(16):3280.
. Scavelli C, Nico B, Cirulli T, et al. Vasculogenic mimicry by bone marrow macrophages in patients with multiple myeloma. Oncogene 2008;27(5):663.
. Chaouchi N, Wallon C, Goujard C, et al. Interleukin-13 inhibits interleukin-2-induced proliferation and protects chronic lymphocytic leukemia B cells from in vitro apoptosis. Blood 1996;87(3):1022.
. DiLillo DJ, Weinberg JB, Yoshizaki A, et al. Chronic lymphocytic leukemia and regulatory B cells share IL-10 competence and immunosuppressive function. Leukemia 2013;27(1):170.
. Chen YCE, Mapp S, Blumenthal A, et al. The duality of macrophage function in chronic lymphocytic leukaemia. Biochim Biophys Acta Rev Cancer 2017;1868(1):176.
. Galletti G, Scielzo C, Barbaglio F, et al. Targeting macrophages sensitizes chronic lymphocytic leukemia to apoptosis and inhibits disease progression. Cell Rep 2016;14(7):1748.
. Blunt MD, Carter MJ, Larrayoz M, et al. The PI3K/mTOR inhibitor PF-04691502 induces apoptosis and inhibits microenvironmental signaling in CLL and the Emicro-TCL1 mouse model. Blood 2015;125(26):4032.
. Petty AJ, Yang Y. Tumor-associated macrophages: implications in cancer immunotherapy. Immunotherapy 2017;9(3):289.
. Franklin RA, Liao W, Sarkar A, et al. The cellular and molecular origin of tumor-associated macrophages. Science 2014;344(6186):921.
. Movahedi K, Laoui D, Gysemans C, et al. Different tumor microenvironments contain functionally distinct subsets of macrophages derived from Ly6C(high) monocytes. Cancer Res 2010;70(14):5728.
. Komohara Y, Fujiwara Y, Ohnishi K, Takeya M. Tumor-associated macrophages: potential therapeutic targets for anti-cancer therapy. Adv Drug Deliv Rev 2016;99(Pt B):180.
. Ngambenjawong C, Gustafson HH, Pun SH. Progress in tumor-associated macrophage (TAM)-targeted therapeutics. Adv Drug Deliv Rev 2017;114:206.
. Pathria P, Louis TL, Varner JA. Targeting tumor-associated macrophages in cancer. Trends Immunol 2019;40(4):310.
. Biswas SK, Mantovani A. Macrophage plasticity and interaction with lymphocyte subsets: cancer as a paradigm. Nat Immunol 2010;11(10):889.
. Ries CH, Cannarile MA, Hoves S, et al. Targeting tumor-associated macrophages with anti-CSF-1R antibody reveals a strategy for cancer therapy. Cancer Cell 2014;25(6):846.
. Hanna BS, McClanahan F, Yazdanparast H, et al. Depletion of CLL-associated patrolling monocytes and macrophages controls disease development and repairs immune dysfunction in vivo. Leukemia 2016;30(3):570.
. Theocharides AP, Jin L, Cheng PY, et al. Disruption of SIRPα signaling in macrophages eliminates human acute myeloid leukemia stem cells in xenografts. J Exp Med 2012;209(10):1883.
. Morrissey MA, Williamson AP, Steinbach AM, et al. Chimeric antigen receptors that trigger phagocytosis. Elife 2018;7.
. Schulz C, Gomez Perdiguero E, Chorro L, et al. A lineage of myeloid cells independent of Myb and hematopoietic stem cells. Science 2012;336(6077):86.
. Gautier EL, Shay T, Miller J, et al. Gene-expression profiles and transcriptional regulatory pathways that underlie the identity and diversity of mouse tissue macrophages. Nat Immunol 2012;13(11):1118.
. Davies LC, Jenkins SJ, Allen JE, Taylor PR. Tissue-resident macrophages. Nat Immunol 2013;14(10):986.
. Mass E, Ballesteros I, Farlik M, et al. Specification of tissue-resident macrophages during organogenesis. Science 2016;353(6304):
. Prinz M, Priller J, Sisodia SS, Ransohoff RM. Heterogeneity of CNS myeloid cells and their roles in neurodegeneration. Nat Neurosci 2011;14(10):1227.
. Taylor PR, Brown GD, Reid DM, et al. The beta-glucan receptor, dectin-1, is predominantly expressed on the surface of cells of the monocyte/macrophage and neutrophil lineages. J Immunol 2002;169(7):3876.
. Hashimoto D, Miller J, Merad M. Dendritic cell and macrophage heterogeneity in vivo. Immunity 2011;35(3):323.
. Chorro L, Sarde A, Li M, et al. Langerhans cell (LC) proliferation mediates neonatal development, homeostasis, and inflammation-associated expansion of the epidermal LC network. J Exp Med 2009;206(13):3089.
. Klein I, Cornejo JC, Polakos NK, et al. Kupffer cell heterogeneity: functional properties of bone marrow derived and sessile hepatic macrophages. Blood 2007;110(12):4077.
. Sadahira Y, Mori M. Role of the macrophage in erythropoiesis. Pathol Int 1999;49(10):841.