IL1R2 Blockade Alleviates Immunosuppression and Potentiates Anti-PD-1 Efficacy in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with limited therapeutic options. IL1 receptor type 2 (IL1R2) promotes breast tumor-initiating cell (BTIC) self-renewal and tumor growth in TNBC, indicating that targeting it could improve patient treatment. In this study, we observed that IL1R2 blockade strongly attenuated macrophage recruitment and the polarization of tumor-associated macrophages (TAM) to inhibit BTIC self-renewal and CD8+ T-cell exhaustion, which resulted in reduced tumor burden and prolonged survival in TNBC mouse models. IL1R2 activation by TAM-derived IL1ß increased PD-L1 expression by interacting with the transcription factor Yin Yang 1 (YY1) and inducing YY1 ubiquitination and proteasomal degradation in both TAMs and TNBC cells. Loss of YY1 alleviated the transcriptional repression of c-Fos, which is a transcriptional activator of PDL-1. Combined treatment with an IL1R2-neutralizing antibodies and anti-PD-1 led to enhanced antitumor efficacy and reduced TAMs, BTICs, and exhausted CD8+ T cells. These results suggest that IL1R2 blockade might be a strategy to potentiate immune checkpoint blockade efficacy in TNBC to improve patient outcomes. Significance: IL1R2 in both macrophages and breast cancer cells orchestrates an immunosuppressive tumor microenvironment by upregulating PD-L1 expression and can be targeted to enhance the efficacy of anti-PD-1 in triple-negative breast cancer.

Superparamagnetic Iron Oxide Nanoparticles Reprogram the Tumor Microenvironment and Reduce Lung Cancer Regrowth after Crizotinib Treatment.

ALK-positive NSCLC patients demonstrate initial responses to ALK tyrosine kinase inhibitor (TKI) treatments, but eventually develop resistance, causing rapid tumor relapse and poor survival rates. Growing evidence suggests that the combination of drug and immune therapies greatly improves patient survival; however, due to the low immunogenicity of the tumors, ALK-positive patients do not respond to currently available immunotherapies. Tumor-associated macrophages (TAMs) play a crucial role in facilitating lung cancer growth by suppressing tumoricidal immune activation and absorbing chemotherapeutics. However, they can also be programmed toward a pro-inflammatory tumor suppressive phenotype, which represents a highly active area of therapy development. Iron loading of TAMs can achieve such reprogramming correlating with an improved prognosis in lung cancer patients. We previously showed that superparamagnetic iron oxide nanoparticles containing core-cross-linked polymer micelles (SPION-CCPMs) target macrophages and stimulate pro-inflammatory activation. Here, we show that SPION-CCPMs stimulate TAMs to secrete reactive nitrogen species and cytokines that exert tumoricidal activity. We further show that SPION-CCPMs reshape the immunosuppressive Eml4-Alk lung tumor microenvironment (TME) toward a cytotoxic profile hallmarked by the recruitment of CD8+ T cells, suggesting a multifactorial benefit of SPION-CCPM application. When intratracheally instilled into lung cancer-bearing mice, SPION-CCPMs delay tumor growth and, after first line therapy with a TKI, halt the regrowth of relapsing tumors. These findings identify SPIONs-CCPMs as an adjuvant therapy, which remodels the TME, resulting in a delay in the appearance of resistant tumors.

Arenobufagin modulation of PCSK9-mediated cholesterol metabolism induces tumor-associated macrophages polarisation to inhibit hepatocellular carcinoma progression.

BACKGROUND: The tumor microenvironment (TME) of hepatocellular carcinoma is heterogeneous enough to be prone to drug resistance and multidrug resistance during treatment, and reprogramming of cholesterol metabolism in TME mediates tumor-associated macrophages (TAMs) polarization, which has an impact on the regulation of malignant tumor progression. Arenobufagin (ARBU) was extracted and isolated from toad venom (purity ≥98 %), which is the main active ingredient of the traditional Chinese medicine Chan'su with good anti-tumor effects. PURPOSE: To investigate the regulatory effect of ARBU on lipid metabolism in tumor microenvironment, interfere with macrophage polarization, and determine its mechanism of action on liver cancer progression. METHODS: In this study, the inhibitory effect of ARBU on the proliferation of Hepa1-6 in C57 mice and the safety of administration were evaluated by establishing a transplanted tumor model of Hepa1-6 hepatocellular carcinoma mice and using 5-FU as a positive control drug. In addition, we constructed a co-culture system of Hepa1-6 cells and primary mouse macrophages to study the effects of ARBU on the polarization phenotypic transformation of macrophages and the proliferation and migration of hepatoma cells. The influence of ARBU on the metabolism of lipids in the hepatocellular carcinoma mouse model was investigated by combining it with lipidomics technology. The influence of ARBU on the PCSK9/LDL-R signaling pathway and macrophage polarization, which regulate cholesterol metabolism, was tested by using qRT-PCR, gene editing, IF, and WB. CONCLUSION: ARBU significantly inhibited the proliferation of Hepa1-6 in vivo and in vitro, regulated cholesterol metabolism, and promoted the M1-type polarization of macrophages in the tumor microenvironment. ARBU inhibits cholesterol synthesis in the TME through the PCSK9/LDL-R signaling pathway, thereby blocking macrophage M2 polarization, promoting apoptosis of the tumor cells, and inhibiting their proliferation and migration.

Calcitriol promotes M2 polarization of tumor-associated macrophages in 4T1 mouse mammary gland cancer via the induction of proinflammatory cytokines.

Our research found that vitamin D3 (VD3) treatment increased lung metastasis in mice with 4T1 murine breast cancer (BC). This study aims to investigate the impact of VD3 on the activation of tumor-associated macrophages (TAMs) in BC. Mice bearing 4T1, E0771, 67NR BC cells, and healthy mice, were fed diets with varying VD3 contents (100-deficient, 1000-normal, and 5000 IU/kg-elevated). Some mice in the 1000 and 100 IU/kg groups received calcitriol. We studied bone metastasis and characterized TAMs and bone marrow-derived macrophages (BMDMs). 4T1 cells had higher bone metastasis potential in the 5000 IU/kg and calcitriol groups. In the same mice, an elevated tumor osteopontin level and M2 polarization of TAMs (MHCIIlow CD44high phenotype) were observed. Gene expression analysis confirmed M2 polarization of 4T1 (but not 67NR) TAMs and BMDMs, particularly in the 100 IU + cal group (increased Mrc1, Il23, and Il6). This polarization was likely due to COX-2/PGE2 induction in 4T1 calcitriol-treated cells, leading to increased proinflammatory cytokines like IL-6 and IL-23. Future studies will explore COX-2/PGE2 as a primary mediator of calcitriol-stimulated inflammation in the BC microenvironment, especially relevant for BC patients with VD3 deficiency and supplementation.

Tumor-associated macrophages in colorectal cancer metastasis: molecular insights and translational perspectives.

Metastasis is the leading cause of high mortality in colorectal cancer (CRC), which is not only driven by changes occurring within the tumor cells, but is also influenced by the dynamic interaction between cancer cells and components in the tumor microenvironment (TME). Currently, the exploration of TME remodeling and its impact on CRC metastasis has attracted increasing attention owing to its potential to uncover novel therapeutic avenues. Noteworthy, emerging studies suggested that tumor-associated macrophages (TAMs) within the TME played important roles in CRC metastasis by secreting a variety of cytokines, chemokines, growth factors and proteases. Moreover, TAMs are often associated with poor prognosis and drug resistance, making them promising targets for CRC therapy. Given the prognostic and clinical value of TAMs, this review provides an updated overview on the origin, polarization and function of TAMs, and discusses the mechanisms by which TAMs promote the metastatic cascade of CRC. Potential TAM-targeting techniques for personalized theranostics of metastatic CRC are emphasized. Finally, future perspectives and challenges for translational applications of TAMs in CRC development and metastasis are proposed to help develop novel TAM-based strategies for CRC precision medicine and holistic healthcare.

Specific molecular weight of Lycium barbarum polysaccharide for robust breast cancer regression by repolarizing tumor-associated macrophages.

The pro-tumorigenic M2-type tumor-associated macrophages (TAMs) in the immunosuppressive tumor microenvironment (TME) promote the progression, angiogenesis, and metastasis of breast cancer. The repolarization of TAMs from an M2-type toward an M1-type holds great potential for the inhibition of breast cancer. Here, we report that Lycium barbarum polysaccharides (LBPs) can significantly reconstruct the TME by modulating the function of TAMs. Specifically, we separated four distinct molecular weight segments of LBPs and compared their repolarization effects on TAMs in TME. The results showed that LBP segments within 50-100 kDa molecular weight range exhibited the prime effect on the macrophage repolarization, augmented phagocytosis effect of the repolarized macrophages on breast cancer cells, and regression of breast tumor in a tumor-bearing mouse model. In addition, RNA-sequencing confirms that this segment of LBP displays an enhanced anti-breast cancer effect through innate immune responses. This study highlights the therapeutic potential of LBP segments within the 50-100 kDa molecular weight range for macrophage repolarization, paving ways to offer new strategies for the treatment of breast cancer.

[Tongxie Yaofang regulates tumor-associated macrophage polarization in colorectal cancer under chronic stress].

This study aims to investigate the intervention effect and mechanism of Tongxie Yaofang in regulating tumor-associated macrophage polarization on colorectal cancer under chronic stress. BALB/C mice were randomized into blank, control, model, mifepristone, and low-, medium-, and high-dose Tongxie Yaofang groups. The other groups except the blank and model groups were subjected to chronic restraint stress and subcutaneous implantation of colon cancer cells for the modeling of colon cancer under stress. Du-ring this period, the body mass and tumor size of each group of mice were recorded. The degree of depression in mice was assessed by behavioral changes. Enzyme-linked immunosorbent assay was employed to determine the levels of cortisol(CORT), 5-hydroxytryptamine(5-HT), norepinephrine(NE), M1-associated inflammatory cytokines [interleukin(IL)-1ß, IL-12, and tumor necrosis factor(TNF)-α], and M2-associated inflammatory cytokines(IL-4 and IL-10) in the serum. The tumor growth of mice in each group was regularly monitored by in vivo imaging. The histopathological changes of tumors in each group of mice were observed by hematoxylin-eosin staining. The proportions of CD86 and CD206 in the tumor tissue were detected by flow cytometry and immunofluorescence staining. Western blot was employed to determine the protein levels of Janus kinase(JAK)1, JAK2, JAK3, signal transducer and activator of transcription(STAT)3, and STAT6 in the tumor tissue. The results showed that chronic stress increased the immobility time of mice, elevated the serum levels of CORT, IL-4, and IL-10, lowered the levels of 5-HT, NE, IL-1ß, IL-12, and TNF-α, and promoted the growth of subcutaneous tumors. The tumor cells in the tumor tissue grew actively, with obvious atypia and up-regulated protein levels of CD206, JAK1, JAK2, JAK3, STAT3, and STAT6, and down-regulated protein level of CD86. The treatment with Tongxie Yaofang shortened the immobility time of mice, lowered the serum levels of CORT, IL-4, and IL-10, elevated the serum levels of 5-HT, NE, IL-1ß, IL-12, and TNF-α, and inhibited the growth of subcutaneous tumors in mice. Moreover, the treatment caused different degrees of necrosis in the tumor tissues, down-regulated the protein levels of CD206, JAK1, JAK2, JAK3, STAT3, and STAT6, and up-regulated the protein level of CD86. In summary, Tongxie Yaofang can promote the transformation of M2 macrophages to M1 macrophages and change the tumor microenvironment under chronic stress to inhibit the development of colorectal cancer, which may be related to the JAK/STAT signaling pathway.

Blocking LTB4 signaling-mediated TAMs recruitment by Rhizoma Coptidis sensitizes lung cancer to immunotherapy.

BACKGROUND: Immune checkpoint blockade (ICB) induces durable immune responses across a spectrum of advanced cancers and revolutionizes the oncology field. However, only a subset of patients achieves long-lasting clinical benefits. Tumor-associated macrophages (TAMs) usually secrete immunosuppressive cytokines and contribute to the failure of ICB therapy. Therefore, it is crucial to mechanically manipulate the abundance and function of TAMs in the tumor microenvironment (TME), which can offer a promising molecular basis to improve the clinical response efficacy of ICB in cancer patients. PURPOSE: This study aims to investigate TAMs in the immunosuppressive microenvironment to identify new therapeutic targets, improve the ability to predict and guide responses to clinical immunotherapy, and develop new strategies for immunotherapy of lung tumors. METHODS: Lewis lung carcinoma (LLC) xenograft-bearing mouse models were established to analyze the antitumor activity of Rhizoma Coptidis (RC) in vivo. A systems pharmacology strategy was used to predict the correlation between RC and M2 macrophages. The effect of RC on the abundance of M2 macrophages was analyzed by flow cytometry of murine samples. Western blot was performed to analyze the expression of Leukotriene A4 hydrolase (LTA4H) and LTB4 receptor 1 (BLT1) in harvested lung cancer tissues. The impact of blocking leukotriene B4 (LTB4) signaling by RC on the recruitment of M2 macrophages was assessed in vitro and in vivo. Transwell migration assays were conducted to clarify the inhibition of macrophage migration by blocking LTB4. Lta4h-/- mice were used to investigate the sensitivity of immunotherapy to lung cancer by blocking the LTB4 signaling. RESULTS: Here, we report that RC, an herbal medicine from the family Ranunculaceae, suppresses the recruitment and immunosuppressive function of TAMs, which in turn sensitizes lung cancer to ICB therapy. Firstly, a systems pharmacology strategy was proposed to identify combinatorial drugs for ICB therapy with a systems biology perspective of drug-target-pathway-TME phenotype. We predicted and verified that RC significantly inhibits tumor growth and the infiltration of M2-TAMs into TME of LLC tumor-bearing mice. Then, RC inhibits the recruitment of macrophages to the tumor TME via blocking LTB4 signaling, and suppresses the expression of immunosuppressive factors (IL-10, TGF-ß and VEGF). As a result, RC enables CD8+ T cells to retain their proliferative and infiltrative abilities within the TME. Ultimately, these events promote cytotoxic T-cell-mediated clearance of tumor cells, which is further enhanced by the addition of anti-PD-L1 therapy. Furthermore, we employed LTA4H deficient mice (Lta4h-/- mice) to evaluate the antitumor efficiency, the results showed that the efficacy of immunotherapy was enhanced due to the synergistic effect of LTB4 signaling blockage and ICB inhibition, leading to remarkable inhibition of tumor growth in a mouse model of lung adenocarcinoma. CONCLUSIONS: Taken together, these findings suggest that RC enhances antitumor immunity, providing a rationale for combining RC with immunotherapies as a potential anti-cancer treatment strategy.

[Prevention and treatment of lung cancer by regulating tumor-associated macrophages with traditional Chinese medicine].

Lung cancer is one of the common malignant tumors in the world, and its incidence and mortality is increasing year by year. Interactions between tumor cells and immune cells in the tumor microenvironment(TME) affect tumor proliferation, infiltration, and metastasis. Tumor-associated macrophages(TAMs) are prominent components of TME, and they have dual regulation effects on malignant progression of lung cancer. The number, activity, and function of M2 macrophages are related to the poor prognosis of lung cancer, and M2 macrophages participate in tumor angiogenesis and immune escape. It has been proved that traditional Chinese medicines(TCMs) and their active ingredients can enhance the antitumor effects, reduce the toxicity of chemotherapy and radiotherapy, and prolong the survival rates of patients with cancer. This paper summarized the role of TAMs in the lung cancer initiation and progression, explored the molecular mechanism of TCM in regulating the recruitment, polarization phenotype, activity, and expression of related factors and proteins of TAMs, and discussed related signal pathways in the prevention and treatment of lung cancer based on the TCM theory of "reinforcing healthy qi and eliminating pathogen". This paper is expected to provide new ideas for the immunotherapy of targeted TAMs.

Salvia mitiorrhiza Bunge aqueous extract attenuates infiltration of tumor-associated macrophages and potentiates anti-PD-L1 immunotherapy in colorectal cancer through modulating Cox2/PGE2 cascade.

ETHNOPHARMACOLOGICAL RELEVANCE: Based on the notion of traditional Chinese medicine, the theory of invigorating the circulation of blood is a prominent treatment for cancer in clinic. Therefore, Salvia miltiorrhiza Bunge, as a representative of Chinese medicine of invigorating the circulation of blood, has been proved to be an effective medicinal herb for treating cancer. AIM OF THE STUDY: To clarify the anti-cancer effect of Salvia miltiorrhiza Bunge aqueous extract (SMAE) on colorectal cancer (CRC) and investigate whether the therapeutic effect of SMAE was mediated by attenuating the infiltration of tumor-associated macrophages (TAMs) into the tumor microenvironment (TME). MATERIALS AND METHODS: High-performance liquid chromatography (HPLC) was used for determined the main compounds of SMAE. MC38 cells were subcutaneously injected into the mice to establish the mouse model of CRC. Tumor growth curve was detected by tumor volume measurement. The model group received distilled water irrigation once a day. SMAE-treated group received 5 g/kg or 10 g/kg SMAE once a day. Anti-PD-L1 treated group received 5 mg/kg anti-PD-L1 once every three days. Protein expression of Cox2 and PD-L1 was determined by Western blot assay. The secretion levels of PGE2, IL-1ß, IL-6, MCP-1, and GM-CSF were evaluated through ELISA. The mRNA expression of CSF1, CCL2, CXCL1, CXCL2, and CXCL3 was measured by using RT-qPCR. Staining of Ki67, TUNEL and Caspase3 was used to investigate cell proliferation and apoptosis. Immunohistochemical staining was used to determine CD8+ T cell distribution. H&E staining was used to confirm histopathological changes. The expressions of F4/80 and CD68 were measured by flow cytometry to identify macrophages in tumors and lymph nodes. The number of CD8+ T cells and the expression of PD-1, IFN-γ, and Granzyme B (GZMB) were determined by flow cytometry. RESULTS: SMAE significantly retarded the growth of MC38 mouse colorectal cancer. SMAE strikingly inhibited the expression of Cox2 and impaired the secretion of PGE2 in tumors, contributing to the attenuated intra-tumoral infiltration of TAMs via Cox2/PGE2 cascade. Meanwhile, SMAE augmented anti-tumor immunity by the elevated proportion of IFN-γ+ CD8+ T cells and GZMB+ CD8+ T cells, which decreased the tumor load. Furthermore, the combination of SMAE and anti-PD-L1 showed a higher therapeutic efficacy than either monotherapy in controlling tumor growth in MC38 xenograft model. CONCLUSIONS: SMAE attenuated the infiltration of TAMs into tumors and synergized with anti-PD-L1 to treat CRC via modulating Cox2/PGE2 cascade.

Garcinone E suppresses breast cancer growth and metastasis by modulating tumor-associated macrophages polarization via STAT6 signaling.

Cancer metastasis remains the most common cause of death in breast cancer patients. Tumor-associated macrophages (TAMs) are a novel therapeutic target for the treatment of metastatic breast cancer. Despite the good anti-cancer activity of garcinone E (GE), there are no reports on its therapeutic effects on breast cancer metastasis. The objective of this study was to examine the anti-cancer effects of GE on metastatic breast cancer. RAW 264.7 and THP-1 cells were polarized to M2 macrophages by IL-4/IL-13 in vitro. A 4T1 mouse breast cancer model and the tail vein breast cancer metastasis model were used to explore the effect of GE on breast cancer growth and metastasis in vivo. In vitro studies showed that GE dose-dependently suppressed IL-4 + IL-13-induced expression of CD206 in both RAW 264.7 cells and differentiated THP-1 macrophages. However, GE did not affect the LPS + IFN-γ-induced polarization to the M1-like macrophages in vitro. GE inhibited the expression of the M2 macrophage specific genes in RAW 264.7 cells, and simultaneously impaired M2 macrophage-induced breast cancer cell proliferation and migration, and angiogenesis. In animal studies, GE significantly suppressed tumor growth, angiogenesis, and lung metastasis in 4T1 tumor-bearing mice, without causing toxicity. In both tumor and lung tissues, the proportion of M2-like TAMs was significantly decreased while the proportion of M1-like TAMs was markedly increased by GE treatment. Mechanistically, GE inhibited phosphorylation of STAT6 in vitro and in vivo. Our results demonstrate for the first time that GE suppresses breast cancer growth and pulmonary metastasis by modulating M2-like macrophage polarization through the STAT6 signaling pathway.

Tetrastigma polysaccharide reprogramming of tumor-associated macrophages via PPARγ signaling pathway to play antitumor activity in breast cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: Tetrastigma Hemsleyanum Diels et Gilg (SYQ) is a typical She ethnomedicine that has been used in anti-tumor treatment in Chinese folklore. The polysaccharide of SYQ (SYQ-PA) has been reported to have antioxidant and anti-inflammatory effects, but the effect and mechanism on antitumor is still unclear. AIM OF THE STUDY: To investigate the activity and mechanism of SYQ-PA against breast cancer in vitro and in vivo. MATERIALS AND METHODS: In this study, different stages of MMTV-PYMT mice, which at 4-week-old and 8-week-old representative the transition from hyperplasia to late carcinoma, were used to investigate the potential effect of SYQ-PA of breast cancer development in vivo. The mechanism was explored with IL4/13-induced peritoneal macrophages model. Flow cytometry assay was employed to analysis the change of tumor microenvironment and the macrophages typing. The inhibition of the condition medium from macrophages on breast cancer cells was detected with xCELLigence system detection. The inflammation factors were tested with cytometric bead array. Co-culture system was used to detect the cell migration and invasion. In addition, the underlying mechanism was investigated using RNAseq analysis, Q-PCR and Western blot, and the PPARγ inhibitor was used to verify the mechanism. RESULTS: SYQ-PA significantly attenuated the process of breast primary tumor growth and reduced the infiltration of TAMs accompanied promoting the polarization of M1 phenotype in MMTV-PyMT mice. Then in vitro studies showed that SYQ-PA promoted macrophages polarization form IL4/13 induced M2 toward to the anti-tumor M1 phenotypes, and the conditioned medium (CM) from the induced macrophages inhibited the proliferation of breast cancer cells. At the same time, SYQ-PA treated macrophages inhibited the migration and invasion of 4T1 in the co-culture system. Further results indicated that SYQ-PA suppressed the release of anti-inflammatory factors and promoted the production of inflammatory cytokines which may induce M1 macrophage polarization and inhibit breast cancer cell proliferation. Subsequently, the underlying mechanism analysis based on RNAseq and molecular assays indicated that SYQ-PA inhibited PPARγ expression and regulated downstream NF-κB in macrophages. After treated with PPARγ inhibitor, T0070907, the effect of SYQ-PA was decreased, or even disappeared. As the downstream, the expression of ß-catenin was also inhibited obviously, those above all contribute the process of SYQ-PA induced M1 macrophages polarization. CONCLUSIONS: Collectively, SYQ-PA was observed inhibited breast cancer, at least in part, via PPARγ activation- and ß-catenin-mediated M2 macrophages polarization. These data expound the antitumor effect and mechanism of SYQ-PA, and provide a possible that SYQ-PA can be used as an adjuvant drug for macrophage tumor immunotherapy in breast cancer.

Crude polysaccharide from Danggui Buxue decoction enhanced the anti-tumor effect of gemcitabine by remodeling tumor-associated macrophages.

Tumor-associated macrophages (TAMs) with an M2-phenotype mediate gemcitabine resistance to cancer by influencing the metabolic enzymes of gemcitabine and releasing competitive deoxycytidine (dC). Our previous studies showed that Danggui Buxue Decoction (DBD), a traditional Chinese medicinal recipe, enhances the anti-tumor activity of gemcitabine in vivo and alleviates gemcitabine-induced myelosuppression. However, the material basis and exact mechanism underlying its enhanced effects remain unclear. In this study, a bioactive polysaccharide consisting of arabinose, mannose, ribose, and glucose was isolated from DBD. In vivo results demonstrated that DBD crude polysaccharide (DBDP) ameliorated gemcitabine-induced immune system disorders. Moreover, DBDP improved the sensitivity of Lewis lung carcinoma-bearing mice to gemcitabine by reshaping the tumor-promoting M2-like macrophages into tumor-inhibiting M1-phenotypes. Furthermore, in vitro results further revealed that DBDP blocked the protective effects of TAMs and M2-macrophages against gemcitabine by inhibiting the excessive secretion of dC and decreasing the high expression of cytidine deaminase. In conclusion, our results demonstrated that DBDP, as the pharmacodynamic material basis of DBD, enhanced the anti-tumor activity of gemcitabine against lung cancer in vitro and in vivo, which was associated with remodeling of the M2-phenotype.

Medicinal plant-derived mtDNA via nanovesicles induces the cGAS-STING pathway to remold tumor-associated macrophages for tumor regression.

Plant-derived nanovesicles (PDNVs) have been proposed as a major mechanism for the inter-kingdom interaction and communication, but the effector components enclosed in the vesicles and the mechanisms involved are largely unknown. The plant Artemisia annua is known as an anti-malaria agent that also exhibits a wide range of biological activities including the immunoregulatory and anti-tumor properties with the mechanisms to be further addressed. Here, we isolated and purified the exosome-like particles from A. annua, which were characterized by nano-scaled and membrane-bound shape and hence termed artemisia-derived nanovesicles (ADNVs). Remarkably, the vesicles demonstrated to inhibit tumor growth and boost anti-tumor immunity in a mouse model of lung cancer, primarily through remolding the tumor microenvironment and reprogramming tumor-associated macrophages (TAMs). We identified plant-derived mitochondrial DNA (mtDNA), upon internalized into TAMs via the vesicles, as a major effector molecule to induce the cGAS-STING pathway driving the shift of pro-tumor macrophages to anti-tumor phenotype. Furthermore, our data showed that administration of ADNVs greatly improved the efficacy of PD-L1 inhibitor, a prototypic immune checkpoint inhibitor, in tumor-bearing mice. Together, the present study, for the first time, to our knowledge, unravels an inter-kingdom interaction wherein the medical plant-derived mtDNA, via the nanovesicles, induces the immunostimulatory signaling in mammalian immune cells for resetting anti-tumor immunity and promoting tumor eradication.

A biomimetic nanoplatform for precise reprogramming of tumor-associated macrophages and NIR-II mediated antitumor immune activation.

The therapeutic effects of photothermal therapy (PTT) are dependent on the photothermal conversion efficiency of photothermal agents (PTAs) in tumors and the subsequent activation of the antitumor immune system. However, the insufficient tumor accumulation of current PTAs and the inevitable recruitment of tumor-associated macrophages (TAMs) could further compromise the antitumor activities of PTT. To address these issues, a biomimetic photothermal nanoplatform Au@Fe-PM is developed for the targeted remodeling of TAMs, which promotes the antitumor immunity of PTT. Au nanorods with second near-infrared (NIR-II) absorptions are fabricated to serve as PTAs to induce immunogenic cell death in tumor cells. The ferric hydroxide shell coated on Au nanorods can release iron ions to repolarize M2-like TAMs into the tumoricidal M1 phenotype via P38 and STAT1-mediated signaling pathways. Moreover, the surface decoration of platelet membranes endows biomimetic nanoplatform with enhanced tumor targeting ability for precise tumor ablation and TAM regulation. Consequently, Au@Fe-PM under NIR-II laser irradiation exhibits significantly higher inhibitory effects in a poor immunogenic 4T1 tumor-bearing mouse model with a 50% complete remission rate compared to conventional PTT (0%). By simultaneously reversing the immunosuppressive tumor microenvironment, this biomimetic nanoplatform offers a promising strategy for enhancing the antitumor efficacy of PTT. STATEMENT OF SIGNIFICANCE: The therapeutic effects of current photothermal therapy (PTT) are hindered by the insufficient tumor accumulation of conventional photothermal agents and the recruitment of immunosuppressive tumor-associated macrophages (TAMs) after PTT. Herein, we report a biomimetic iron-based second near-infrared (NIR-II) photothermal nanoplatform (Au@Fe-PM) for targeted TAMs reprogramming and NIR-II mediated anti-tumor immunity. Au@Fe-PM can actively target the tumor site with the help of surface-decorated platelet membranes. Meanwhile, iron ions would be released from Au@Fe-PM in acidic lysosomes to reprogram TAMs into tumoricidal M1-like macrophages, which promotes the antitumor responses elicited by NIR-II PTT, thereby contributing to remarkable tumor inhibitory effects, with 50% higher complete remission rate than that of conventional PTT.

7S,15R-Dihydroxy-16S,17S-epoxy-docosapentaenoic Acid Overcomes Chemoresistance of 5-Fluorouracil by Suppressing the Infiltration of Tumor-Associated Macrophages and Inhibiting the Activation of Cancer Stem Cells in a Colorectal Cancer Xenograft Model.

Although the tumor bulk is initially reduced by 5-fluorouracil (5-FU), chemoresistance developed due to prolonged chemotherapy in colorectal cancer (CRC). The enrichment of cancer stem cells (CSCs) and the infiltration of tumor-associated macrophages (TAMs) contribute to chemoresistance and poor outcomes. A docosahexaenoic acid derivative developed by our group, 7S,15R-dihydroxy-16S,17S-epoxy-docosapentaenoic acid (diHEP-DPA), exerts antitumor effects against TAMs infiltration and CSCs enrichment in our previous study. The current study aimed to investigate whether diHEP-DPA was able to overcome chemoresistance to 5-FU in CRCs, together with the potential synergistic mechanisms in a CT26-BALB/c mouse model. Our results suggested that although 5-FU inhibited tumor growth, 5-FU enriched CSCs via the WNT/ß-catenin signaling pathway, resulting in chemoresistance in CRCs. However, we revealed that 5-FU promoted the infiltration of TAMs via the NF-kB signaling pathway and improved epithelial-mesenchymal transition (EMT) via the signal transducer and activator of the transcription 3 (STAT3) signaling pathway; these traits were believed to contribute to CSC activation. Furthermore, supplementation with diHEP-DPA could overcome drug resistance by decreasing the CSCs, suppressing the infiltration of TAMs, and inhibiting EMT progression. Additionally, the combinatorial treatment of diHEP-DPA and 5-FU effectively enhanced phagocytosis by blocking the CD47/signal regulatory protein alpha (SIRPα) axis. These findings present that diHEP-DPA is a potential therapeutic supplement to improve drug outcomes and suppress chemoresistance associated with the current 5-FU-based therapies for colorectal cancer.

Integrative analysis of bulk and single-cell gene expression profiles to identify tumor-associated macrophage-derived CCL18 as a therapeutic target of esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is a common gastrointestinal malignancy with poor patient prognosis. Current treatment for ESCC, including immunotherapy, is only beneficial for a small subset of patients. Better characterization of the tumor microenvironment (TME) and the development of novel therapeutic targets are urgently needed. METHODS: In the present study, we hypothesized that integration of single-cell transcriptomic sequencing and large microarray sequencing of ESCC biopsies would reveal the key cell subtypes and therapeutic targets that determine the prognostic and tumorigenesis of ESCC. We characterized the gene expression profiles, gene sets enrichment, and the TME landscape of a microarray cohort including 84 ESCC tumors and their paired peritumor samples. We integrated single-cell transcriptomic sequencing and bulk microarray sequencing of ESCC to reveal key cell subtypes and druggable targets that determine the prognostic and tumorigenesis of ESCC. We then designed and screened a blocking peptide targeting Chemokine C-C motif ligand 18 (CCL18) derived from tumor associated macrophages and validated its potency by MTT assay. The antitumor activity of CCL18 blocking peptide was validated in vivo by using 4-nitroquinoline-1-oxide (4-NQO) induced spontaneous ESCC mouse model. RESULTS: Comparative gene expression and cell-cell interaction analyses revealed dysregulated chemokine and cytokine pathways during ESCC carcinogenesis. TME deconvolution and cell interaction analyses allow us to identify the chemokine CCL18 secreted by tumor associated macrophages could promote tumor cell proliferation via JAK2/STAT3 signaling pathway and lead to poor prognosis of ESCC. The peptide Pep3 could inhibit the proliferation of EC-109 cells promoted by CCL18 and significantly restrain the tumor progression in 4-NQO-induced spontaneous ESCC mouse model. CONCLUSIONS: For the first time, we discovered and validated that CCL18 blockade could significantly prevent ESCC progression. Our study revealed the comprehensive cell-cell interaction network in the TME of ESCC and provided novel therapeutic targets and strategies to ESCC treatment.

Single-cell RNA datasets and bulk RNA datasets analysis demonstrated C1Q+ tumor-associated macrophage as a major and antitumor immune cell population in osteosarcoma.

Background: Osteosarcoma is the most frequent primary bone tumor with a poor prognosis. Immune infiltration proved to have a strong impact on prognosis. We analyzed single-cell datasets and bulk datasets to confirm the main immune cell populations and their properties in osteosarcoma. Methods: The examples in bulk datasets GSE21257 and GSE32981 from the Gene Expression Omnibus database were divided into two immune infiltration level groups, and 34 differentially expressed genes were spotted. Then, we located these genes among nine major cell clusters and their subclusters identified from 99,668 individual cells in single-cell dataset GSE152048 including 11 osteosarcoma patients. Especially, the markers of all kinds of myeloid cells identified in single-cell dataset GSE152048 were set to gene ontology enrichment. We clustered the osteosarcoma samples in the TARGET-OS from the Therapeutically Applicable Research to Generate Effective Treatments dataset into two groups by complete component 1q positive macrophage markers and compared their survival. Results: Compared with the low-immune infiltrated group, the high-immune infiltrated group showed a better prognosis. Almost all the 34 differentially expressed genes expressed higher or exclusively among myeloid cells. A group of complete component 1q-positive macrophages was identified from the myeloid cells. In the bulk dataset TARGET-OS, these markers and the infiltration of complete component 1q-positive macrophages related to longer survival. Conclusions: Complete component 1q-positive tumor-associated macrophages were the major immune cell population in osteosarcoma, which contributed to a better prognosis.

Targeted treatment of triple-negative-breast cancer through pH-triggered tumour associated macrophages using smart theranostic nanoformulations.

Triple-negative breast cancer (TNBC) represents 15-25 % of the new breast cancer cases diagnosed worldwide every year. TNBC is among the most aggressive and worst prognosis breast cancer, mainly because targeted therapies are not available. Herein, we developed a magnetic theranostic hybrid nanovehicle for targeted treatment of TNBC through pH-triggered tumour associated macrophages (TAMs) targeting. The lipid core of the nanovehicle was composed of a Carnaúba wax matrix that simultaneously incorporated iron oxide nanoparticles and doxorubicin (DOX) - a chemotherapeutic drug. These drug-loaded wax nanovehicles were modified with a combination of two functional and complementary molecules: (i) a mannose ligand (macrophage targeting) and (ii) an acid-sensitive sheddable polyethylene glycol (PEG) moiety (specificity). The TAMs targeting strategy relied on the mannose - mannose receptor recognition exclusively after acid-sensitive "shedding" of the PEG in the relatively low tumour microenvironment pH. The pH-induced targeting capability towards TAMs was confirmed in vitro in a J774A.1 macrophage cell line at different pH (7.4 and 6.5). Biocompatibility and efficacy of the final targeted formulations were demonstrated in vitro in the TNBC MDA-MB-231 cell line and in vivo in an M-Wnt tumour-bearing (TNBC) mouse model. A preferential accumulation of the DOX-loaded lipid nanovehicles in the tumours of M-Wnt-tumour bearing mice was observed, which resulted both on an efficient tumour growth inhibition and a significantly reduced off-target toxicity compared to free DOX. Additionally, the developed magnetic hybrid nanovehicles showed outstanding performances as T2-contrast agents in magnetic resonance imaging (r2 ≈ 400-600 mM-1·s-1) and as heat generating sources in magnetic hyperthermia (specific absorption rate, SAR ≈ 178 W·g-1Fe). These targeted magnetic hybrid nanovehicles emerge as a suitable theranostic option that responds to the urgent demand for more precise and personalized treatments, not only because they are able to offer localized imaging and therapeutic potential, but also because they allow to efficiently control the balance between safety and efficacy.

Dioscin induces M1 macrophage polarization through Connexin-43 Channels in Tumor-associated-macrophages-mediated melanoma metastasis.

BACKGROUND: Tumor-associated macrophages (TAMs) are important constituent parts of tumor microenvironment that connected with tumor metastasis in melanoma. Connexin 43 (Cx43) was expressed in all the immune cells which modulated different aspects of immune response. However, the concrete molecular mechanism maintains unclear. PURPOSE: The study aimed to find a natural drug monomer effectively reversed the polarity of tumor-associated macrophages inhibiting melanoma metastasis and improving survival time. METHODS: Flow cytometry was used to determine the effects of dioscin on the macrophage phenotype. Western bolt and ELISA were performed to explore the underlying mechanism of dioscin and a co-culture experiment in vitro was applied to assess the role of dioscin on TAMs-mediated melanoma proliferation, invasion and migration. Moreover, in vivo melanoma metastasis models were established for examining effects of dioscin on TAMs-mediated melanoma metastasis. RESULTS: Dioscin repolarized macrophages from M2 towards M1-like phenotype. Dioscin suppressed M2-like phenotype macrophages through enhanced the expression and transport function of Cx43. Furthermore, the stimulation IFN-γ/STAT1 pathway and suppression IL-4/JAK2/STAT3 pathway were major mechanism of dioscin. Importantly, dioscin suppressed Cx43G21R mutation TAMs induced proliferation, invasion, migration and metastasis of melanoma cells. It worthily noting that dioscin ameliorated tumor-associated-macrophages-mediated melanoma metastasis in vitro and vivo. CONCLUSION: Dioscin re-polarized macrophages from M2 to M1 phenotype through activation of Cx43-gap-junction-intercellular-communications (Cx43-GJs)/IFN-γ/STAT1 pathway and inhibition of Cx43-GJs/IL-4/JAK2/STAT3 suppressing migration, invasion and metastasis of melanoma, which provided a theoretical and experimental basis for treating melanoma metastasis.