The dynamic interactions of cancer cells with their microenvironment consisting of stromal cells (cellular part) and extracellular matrix (ECM) components (non-cellular) is essential to stimulate the heterogeneity of cancer cell, clonal evolution and to increase the multidrug resistance ending in cancer cell progression and metastasis. The reciprocal cell-cell/ECM interaction and tumor cell hijacking of non-malignant cells force stromal cells to lose their function and acquire new phenotypes that promote development and invasion of tumor cells. Understanding the underlying cellular and molecular mechanisms governing these interactions can be used as a novel strategy to indirectly disrupt cancer cell interplay and contribute to the development of efficient and safe therapeutic strategies to fight cancer. Furthermore, the tumor-derived circulating materials can also be used as cancer diagnostic tools to precisely predict and monitor the outcome of therapy. This review evaluates such potentials in various advanced cancer models, with a focus on 3D systems as well as lab-on-chip devices.

https://link.springer.com/content/pdf/10.1186/s12964-020-0530-4.pdf

Tumor microenvironment complexity and therapeutic implications at a glance.

Strategies to therapeutically target the tumor microenvironment (TME) have emerged as a promising approach for cancer treatment in recent years due to the critical roles of the TME in regulating tumor progression and modulating response to standard-of-care therapies Here, we summarize the current knowledge regarding the most advanced TME-directed therapies, which have either been clinically approved or are currently being evaluated in trials, including immunotherapies, antiangiogenic drugs, and treatments directed against cancer-associated fibroblasts and the extracellular matrix We also discuss some of the challenges associated with TME therapies, and future perspectives in this evolving field SIGNIFICANCE: This review provides a comprehensive analysis of the current therapies targeting the TME, combining a discussion of the underlying basic biology with clinical evaluation of different therapeutic approaches, and highlighting the challenges and future perspectives

Therapeutic Targeting of the Tumor Microenvironment

The immunosuppressive status of the tumor microenvironment (TME) remains poorly defined due to a lack of understanding regarding the function of tumor-associated macrophages (TAMs), which are abundant in the TME. TAMs are crucial drivers of tumor progression, metastasis, and resistance to therapy. Intra- and inter-tumoral spatial heterogeneities are potential keys to understanding the relationships between subpopulations of TAMs and their functions. Antitumor M1-like and pro-tumor M2-like TAMs coexist within tumors, and the opposing effects of these M1/M2 subpopulations on tumors directly impact current strategies to improve antitumor immune responses. Recent studies have found significant differences among monocytes or macrophages from distinct tumors, and other investigations have explored the existence of diverse TAM subsets at the molecular level. In this review, we discuss emerging evidence highlighting the redefinition of TAM subpopulations and functions in the TME and the possibility of separating macrophage subsets with distinct functions into antitumor M1-like and pro-tumor M2-like TAMs during the development of tumors. Such redefinition may relate to the differential cellular origin and monocyte and macrophage plasticity or heterogeneity of TAMs, which all potentially impact macrophage biomarkers and our understanding of how the phenotypes of TAMs are dictated by their ontogeny, activation status, and localization. Therefore, the detailed landscape of TAMs must be deciphered with the integration of new technologies, such as multiplexed immunohistochemistry (mIHC), mass cytometry by time-of-flight (CyTOF), single-cell RNA-seq (scRNA-seq), spatial transcriptomics, and systems biology approaches, for analyses of the TME.

/pdf/redefining-tumor-associated-macrophage-subpopulations-and-2kfij88fz3.pdf

Redefining Tumor-Associated Macrophage Subpopulations and Functions in the Tumor Microenvironment.

Exosome-encapsulated miRNAs contribute to CXCL12/CXCR4-induced liver metastasis of colorectal cancer by enhancing M2 polarization of macrophages.

Tumor-associated macrophages (TAMs) are major innate immune cells that constitute up to 50% of the cell mass of human tumors. TAMs are highly heterogeneous cells that originate from resident tissue-specific macrophages and from newly recruited monocytes. TAMs' variability strongly depends on cancer type, stage, and intratumor heterogeneity. Majority of TAMs are programmed by tumor microenvironment to support primary tumor growth and metastatic spread. However, TAMs can also restrict tumor growth and metastasis. In this review, we summarized the knowledge about the role of TAMs in tumor growth, metastasis and in the response to cancer therapy in patients with five aggressive types of cancer: breast, colorectal, lung, ovarian, and prostate cancers that are frequently metastasize into distant organs resulting in high mortality of the patients. Two major TAM parameters are applied for the evaluation of TAM correlation with the cancer progression: total amount of TAMs and specific phenotype of TAMs identified by functional biomarkers. We summarized the data generated in the wide range of international patient cohorts on the correlation of TAMs with clinical and pathological parameters of tumor progression including lymphatic and hematogenous metastasis, recurrence, survival, therapy efficiency. We described currently available biomarkers for TAMs that can be measured in patients' samples (tumor tissue and blood). CD68 is the major biomarker for the quantification of total TAM amounts, while transmembrane receptors (stabilin-1, CD163, CD206, CD204, MARCO) and secreted chitinase-like proteins (YKL-39, YKL-40) are used as biomarkers for the functional TAM polarization. We also considered that specific role of TAMs in tumor progression can depend on the localization in the intratumoral compartments. We have made the conclusion for the role of TAMs in primary tumor growth, metastasis, and therapy sensitivity for breast, colorectal, lung, ovarian, and prostate cancers. In contrast to other cancer types, majority of clinical studies indicate that TAMs in colorectal cancer have protective role for the patient and interfere with primary tumor growth and metastasis. The accumulated data are essential for using TAMs as biomarkers and therapeutic targets to develop cancer-specific immunotherapy and to design efficient combinations of traditional therapy and new immunomodulatory approaches.

/pdf/tumor-associated-macrophages-in-human-breast-colorectal-lung-55u30ctebp.pdf

Tumor-Associated Macrophages in Human Breast, Colorectal, Lung, Ovarian and Prostate Cancers

Tumor-associated macrophages (TAMs) are major components of tumor microenvironment that frequently associated with tumor metastasis in human cancers. Circulating tumor cell (CTC), originating from primary tumor sites, is considered to be the precursors of tumor metastasis. However, the regulatory mechanism of TAMs in CTC-mediated tumor metastasis still remains unclear. Immunohistochemical staining was used to detect the macrophages infiltration (CD68 and CD163), epithelial–mesenchymal transition (EMT) markers (E-cadherin and Vimentin) expression in serial sections of human colorectal cancer (CRC) specimens. Then, the correlations between macrophages infiltration and clinicopathologic features, mesenchymal CTC ratio, and patients’ prognosis were analyzed. A co-culture assay in vitro was used to evaluate the role of TAMs on CRC EMT, migration and invasion, and ELISA, luciferase reporter assay and CHIP were performed to uncover the underlying mechanism. Furthermore, an in vivo model was carried out to confirm the effect of TAMs on mesenchymal CTC-mediated metastasis. Clinically, CD163+ TAMs infiltrated in invasive front was associated with EMT, mesenchymal CTC ratio, and poor prognosis in patients with CRC. CRC–conditioned macrophages regulated EMT program to enhance CRC cells migration and invasion by secreting IL6. TAMs-derived IL6 activated the JAK2/STAT3 pathway, and activated STAT3 transcriptionally inhibited the tumor suppressor miR-506-3p in CRC cells. miR-506-3p, a key miRNA regulating FoxQ1, was downregulated in CRC cells, resulting in increased FoxQ1 expression, which in turn led to the production of CCL2 that promoted macrophage recruitment. Inhibition of CCL2 or IL6 broke this loop and reduced macrophage migration and mesenchymal CTC-mediated metastasis, respectively. Our data indicates that TAMs induce EMT program to enhance CRC migration, invasion, and CTC-mediated metastasis by regulating the JAK2/STAT3/miR-506-3p/FoxQ1 axis, which in turn leads to the production of CCL2 that promote macrophage recruitment, revealing a new cross-talk between immune cells and tumor cells in CRC microenvironment.

/pdf/crosstalk-between-cancer-cells-and-tumor-associated-42mcjqrlj2.pdf

Crosstalk between cancer cells and tumor associated macrophages is required for mesenchymal circulating tumor cell-mediated colorectal cancer metastasis

Tumor microenvironment (TME) is a complex environment containing tumor cells, tumor-associated macrophages (TAMs), interstitial cells, and non-cellular components. Epithelial–mesenchymal transition (EMT), as a major actor in cancer tumorigenicity and metastasis, was involved in the interaction between TAMs and tumor cells. However, the potential mechanisms of EMT and how EMT-programmed tumor cells affect M2-like TAMs still need further exploration. An integrated analysis of nine CRC miRNA expression datasets was performed. Functional assays, including the EdU, clone formation, wound healing, and transwell assays, were used to determine the anticancer role of miR-195-5p in human CRC progression. Furthermore, RNA immunoprecipitation, RNA decay, and dual-luciferase reporter assays were used to determine the mechanism of miR-195-p CRC progression. Then co-culture, migration, and ELISA assays were applied to determine the role of miR-195-5p in macrophage recruitment and alternative polarization. Xenograft mouse models were used to determine the role of miR-195-5p in CRC tumorigenicity and TAM polarization in vivo. An integrated analysis confirmed that miR-195-5p was significantly downregulated in CRC tissues, and patients with a low level of miR-195-5p had significantly shortened overall survival as revealed by the TCGA-COAD dataset. Altered miR-195-5p in colon cancer cells led to distinct changes of proliferation, migration, invasion, and EMT. Mechanistically, miR-195-5p regulated NOTCH2 expression in a post-transcriptional manner by directly binding to 3′-UTR of the Notch2 mRNA. Subsequently, miR-195-5p/NOTCH2 suppressed GATA3-mediated IL-4 secretion in CRC cells and ultimately inhibited M2-like TAM polarization. miR-195-5p may play a vital role in regulating NOTCH2-mediated tumor cell EMT, thereby affecting IL-4-related M2-like TAM polarization in CRC.

/pdf/mir-195-5p-notch2-mediated-emt-modulates-il-4-secretion-in-47h3nhhbyw.pdf

miR-195-5p/NOTCH2-mediated EMT modulates IL-4 secretion in colorectal cancer to affect M2-like TAM polarization

Tumor-associated macrophages (TAMs) in the tumor microenvironment influence tumor initiation, invasion and metastasis. Several studies have shown that Wnt5a is mainly expressed in the tumor stroma, especially in TAMs. However, whether Wnt5a regulates the polarization and biological function of TAMs in colorectal cancer (CRC) is incompletely understood. Immunofluorescence staining was performed to detect CD68 and Wnt5a expression in colorectal tissues from patients (63 CRC specimens VS 20 normal tissues). RT-qPCR, flow cytometry, ELISA and inhibitors were carried out to explore the role of Wnt5a in the polarization of TAMs. Clone formation and transwell assays were performed to determine the effects of Wnt5a–treated macrophages on tumor proliferation, migration and invasion in vitro. Finally, a xenograft model was applied to confirm the effects of Wnt5a+ TAMs on CRC tumorigenesis. We found that high Wnt5a+CD68+/CD68+ TAMs ratio was significantly associated with poor prognosis in CRC patients and Wnt5a+ TAM was an M2-like TAM subtype. Subsequently, we found that Wnt5a induced macrophages to secrete IL-10, which then acted as an autocrine cytokine to induce M2 polarization of these macrophages. IL-10 neutralizing antibody completely reversed the pro-M2 effect of Wnt5a. Mechanistically, the CaKMII-ERK1/2-STAT3 pathway was required for Wnt5a-mediated IL-10 expression in macrophages. Furthermore, Wnt5a-induced M2 macrophages promoted CRC cells proliferation, migration and invasion; knockdown of Wnt5a in TAMs significantly impaired the pro-tumor functions of TAMs. Our data indicate that Wnt5a could induce M2 polarization of TAMs by regulating CaKMII-ERK1/2-STAT3 pathway–mediated IL-10 secretion, ultimately promoting tumor growth and metastasis of CRC.

/pdf/wnt5a-induced-m2-polarization-of-tumor-associated-27v3dl5qtn.pdf

Wnt5a-induced M2 polarization of tumor-associated macrophages via IL-10 promotes colorectal cancer progression

Background: The interaction and crosstalk between tumor-associated macrophages (TAMs) and epithelial-mesenchymal transition (EMT) has been demonstrated to play a critical role in the progression and metastasis of multiple cancers. However, the roles of the M2-polarized TAMs in different tumor location in EMT and prognosis of colorectal cancer (CRC) have not been elucidated. Therefore, the present study was designed to set up a reliable ratio of CD163+/CD68+ to assess M2-polarized TAMs infiltration in the tumor center (TC) and tumor invasive front (TF) and to further evaluate their prognostic value and biological effects on tumor cells in CRC. Methods: TAMs markers (CD68 and CD163) and EMT markers (E-cadherin and Vimentin) expression were evaluated by immunohistochemistry in 81 patients with CRC. Circulating tumor cells (CTCs) of peripheral blood from above patients was also isolated. The correlation of CD163+/CD68+ ratio in different locations, EMT and CTCs counts were further analyses. Kaplan-Meier and the model analyses of univariate Cox proportional hazards were utilized to compare the survival of patients with high CD163+/CD68+ ratio with those with low CD163+/CD68+ ratio. Furthermore, the effects of the M2-polarized TAMs on growth, migration and invasion of CRC cells were explored in vivo and in vitro co-culture system. Results: The results showed that the level of CD163+/CD68+ ratio in TF was significant higher than that in TC, and higher CD163+/CD68+ TF ratio were closely correlated with enhanced lymphovascular invasion, tumor invasion and TNM stage. Interestingly, higher CD163+/CD68+ TF ratio were also significantly associated with EMT program and CTCs counts. Meanwhile, Kaplan-Meier analysis showed that CD163+/CD68+ TF was associated with both recurrence-free survival (RFS) and overall survival (OS) of patients with CRC. Multivariate Cox regression analyses demonstrated that CD163+/CD68+ TF remained an independent prognostic factor for RFS and OS. Further receiver operating characteristic (ROC) curve analysis found that CD163+/CD68+ TF was a better prognosticator compared with CD68+ TF and CD163+ TF for CRC patients. What's more, M2-polarized TAMs secreted TGF-β to facilitate the EMT, growth, proliferation and invasion of CRC cells by in vivo and in vitro experiments. Conclusions: Our studies preliminarily elucidated the prognostic value of CD163+/CD68+ ratio in different tumor locations and the biological functions of M2-polarized TAMs in CRC progression via TGF-β.

/pdf/elevated-cd163-cd68-ratio-at-tumor-invasive-front-is-closely-3ov1vgrxka.pdf

Elevated CD163+/CD68+ Ratio at Tumor Invasive Front is Closely Associated with Aggressive Phenotype and Poor Prognosis in Colorectal Cancer.

Background: M2 macrophages are crucial components of tumor microenvironment that frequently associated with the resistance of therapeutic treatments in human cancers, but their role in the chemosensitivity of colorectal cancer (CRC) to 5-fluorouracil (5-FU) is still obscure. Methods: In our study, we clarified the biological functions of M2 macrophages and their mechanism on the chemosensitivity of CRC cells to 5-FU. Then, we analyzed the correlation between CCL22 and CD68+ and CD163+ tumor-associated macrophages (TAMs), and further elucidated the prognostic value of CCL22 and CD163+ M2 macrophages in clinical CRC samples. Results: M2 macrophages decreased the inhibitory effect of 5-FU on CRC cells migration and invasion by secreting CCL22, and declined the apoptosis induced by 5-FU. Treated with a neutralizing anti-CCL22 antibody destroyed these effects. We further illuminated that M2 macrophages regulated 5-FU resistance of CRC cells through epithelial-mesenchymal transition (EMT) program, PI3K/AKT pathway, and caspase-mediated apoptosis. Clinically, CCL22 was found to have elevated expression in CRC tissue samples, and was positively associated with CD163+ TAMs. Furthermore, the patients with higher CD163+ M2 macrophages and higher expression of CCL22 in CRC tissues had a lower overall survival (OS) rate compared with lower ones. Conclusion: Our findings indicate that M2 macrophage regulated 5-FU-mediated CRC chemoresistance via the EMT program, PI3K/AKT pathway, and caspase-mediated apoptosis by releasing CCL22.

/pdf/m2-macrophages-confer-resistance-to-5-fluorouracil-in-gvyvupv23n.pdf

Xiaobin Lin

Papers

Crosstalk between cancer cells and tumor associated macrophages is required for mesenchymal circulating tumor cell-mediated colorectal cancer metastasis

miR-195-5p/NOTCH2-mediated EMT modulates IL-4 secretion in colorectal cancer to affect M2-like TAM polarization

Wnt5a-induced M2 polarization of tumor-associated macrophages via IL-10 promotes colorectal cancer progression

Elevated CD163+/CD68+ Ratio at Tumor Invasive Front is Closely Associated with Aggressive Phenotype and Poor Prognosis in Colorectal Cancer.

M2 macrophages confer resistance to 5-fluorouracil in colorectal cancer through the activation of CCL22/PI3K/AKT signaling.