M2-type Tumor-associated Macrophages Research Articles

Abstract PO2-08-06: Metformin may improve the outcome of patients with breast cancer and type 2 diabetes mellitus through the effect of tumor immune microenvironment

Abstract Metformin, an anti-diabetic drug, is known to have anti-tumor effects. We examined the outcome of 177 patients with type 2 diabetes mellitus (T2DM) who received surgery for breast cancer. Among them, 49 patients were treated with drugs including metformin. In those patients, recurrence in distant organs was less frequent and postoperative disease-free survival (DFS) tended to be better than those without metformin intake. In patients who received preoperative systemic therapy (PST), rate of pathological complete response (pCR) was significantly more frequent in patients with metformin treatment (p < 0.05). Multiplex immunohistochemical staining of resected tumors revealed that the density of tumor associated macrophages (TAM), especially of CD68(+)CD163(+) M2-type TAM, was significantly lower in tumors with metformin treatment. In contrast, the rate of CD8(+) phenotype in CD3(+) tumor infiltrating lymphocytes (TILs) was significantly higher in metformin group. The results suggest that metformin can change the immune microenvironment from pro-tumorigenic to anti-tumorigenic status, which leads to the favorable outcome of the patients with breast cancer and T2DM. Citation Format: Satomi Shiba, Michiko Harao, Kasumi Ogihara, Takayo Fukuda, Masako Sakuragi, Kitayama Joji, Naohiro Sata. Metformin may improve the outcome of patients with breast cancer and type 2 diabetes mellitus through the effect of tumor immune microenvironment [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-08-06.

Abstract PO1-15-08: Genomic characterization of triple-negative breast cancer metastases reveals PKD1 as a novel biomarker for immunotherapy

Abstract Background: While primary triple-negative breast cancer (TNBC) garners significant research attention, the genomic alterations that occur in metastasis remain insufficiently understood, especially within Asian populations. Furthermore, the genomic information obtained from the primary tumor inadequately guides metastatic cancer treatment, highlighting the critical need for in-depth investigations into metastatic TNBC. Methods: We constructed the largest cohort of TNBC metastases (n = 296) among advanced TNBC patients treated at Fudan University Shanghai Cancer Center (FUSCC) between October 2018 and December 2020. Comprehensive DNA sequencing was conducted on the collected metastatic samples to analyze genomic alterations associated with treatment response. The underlying mechanisms of specific biomarkers were also explored. Results: We presented the genomic landscape of 296 TNBC metastases, encompassing mutant genes, mutation sites and copy number variations. Through multidimensional analysis, significant disparities in TNBC were observed between Western and Asian populations, primary and metastatic tumors, as well as different metastatic sites. Notably, our findings underscore the importance of sequencing TNBC metastases to guide precision therapy, which was associated with longer progression-free survival compared to physician-chosen treatments, shedding light on the pivotal clinical value of genomic studies in metastatic settings. Furthermore, efficacy analysis suggested that PKD1 mutations enriched in metastases mediated resistance to immunotherapy. These findings were further validated through three clinical trials (NCT03805399, NCT04129996, and NCT04395989). Mechanistic studies unveiled the involvement of PKD1 in TNBC immune evasion by upregulating CCL2, thereby facilitating the recruitment of M2-type tumor-associated macrophages. Conclusion: Our study emphasizes the critical significance and necessity of genomic profiling of metastases in guiding precision therapy for TNBC. Moreover, our findings reveal PKD1 as a novel and promising biomarker for immunotherapy. Citation Format: Xiu-Zhi Zhu, Yi-Fan Zhou, Yun-Yi Wang, Xiao-Hong Ding, Xi Jin, Zhi-Ming Shao, Yi-Zhou Jiang, Zhong-Hua Wang. Genomic characterization of triple-negative breast cancer metastases reveals PKD1 as a novel biomarker for immunotherapy [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-15-08.

Optical controlled and nuclear targeted CECR2 competitor to downregulate CSF-1 for metastatic breast cancer immunotherapy

The crosstalk between breast cancer cells and tumor associated macrophages (TAMs) greatly contributes to tumor progression and immunosuppression. In this work, cat eye syndrome chromosome region candidate 2 (CECR2) is identified to overexpress in breast cancer patients, which can recognize v-rel avian reticuloendotheliosis viral oncogene homolog A (RelA) and activate nuclear factor κB (NF-κB) to release colony stimulating factor-1 (CSF-1). Pharmacological inhibition of CECR2 by the bromodomain competitor (Bromosporine, Bro) can downregulate CSF-1 to inhibit M2 type TAMs. To amplify the immunotherapeutic effect, a chimeric peptide-based and optical controlled CECR2 competitor (designated as N-PB) is constructed to enhance the nuclear targeted delivery of Bro and initiate an immunogenic cell death (ICD). In vivo results indicate a favorable breast cancer targeting ability and primary tumor suppression effect of N-PB under optical irradiation. Importantly, N-PB downregulates CSF-1 by competitive inhibition of CECR2 and NF-κB(RelA) interactions, thus inhibiting immunosuppressive M2-like TAMs while improving the antitumorigenic M1-like phenotype. Ultimately, the systemic anti-tumor immunity is activated to suppress the metastatic breast cancer in an optical controlled manner. This study provides a promising therapeutic target and reliable strategy for metastatic breast cancer treatment by interrupting immunosuppressive crosstalk between tumor cells and macrophages.

GPR65 sensing tumor-derived lactate induces HMGB1 release from TAM via the cAMP/PKA/CREB pathway to promote glioma progression

BackgroundLactate has emerged as a critical regulator within the tumor microenvironment, including glioma. However, the precise mechanisms underlying how lactate influences the communication between tumor cells and tumor-associated macrophages (TAMs), the most abundant immune cells in glioma, remain poorly understood. This study aims to elucidate the impact of tumor-derived lactate on TAMs and investigate the regulatory pathways governing TAM-mediated tumor-promotion in glioma.MethodsBioinformatic analysis was conducted using datasets from TCGA and CGGA. Single-cell RNA-seq datasets were analyzed by using UCSC Cell Browser and Single Cell Portal. Cell proliferation and mobility were evaluated through CCK8, colony formation, wound healing, and transwell assays. Western blot and immunofluorescence staining were applied to assess protein expression and cell distribution. RT-PCR and ELISA were employed to identify the potential secretory factors. Mechanistic pathways were explored by western blotting, ELISA, shRNA knockdown, and specific inhibitors and activators. The effects of pathway blockades were further assessed using subcutaneous and intracranial xenograft tumor models in vivo.ResultsElevated expressions of LDHA and MCT1 were observed in glioma and exhibited a positive correlation with M2-type TAM infiltration. Lactate derived from glioma cells induced TAMs towards M2-subtype polarization, subsequently promoting glioma cells proliferation, migration, invasion, and mesenchymal transition. GPR65, highly expressed on TAMs, sensed lactate-stimulation in the TME, fueling glioma cells malignant progression through the secretion of HMGB1. GPR65 on TAMs triggered HMGB1 release in response to lactate stimulation via the cAMP/PKA/CREB signaling pathway. Disrupting this feedback loop by GPR65-knockdown or HMGB1 inhibition mitigated glioma progression in vivo.ConclusionThese findings unveil the intricate interplay between TAMs and tumor cells mediated by lactate and HMGB1, driving tumor progression in glioma. GPR65, selectively highly expressed on TAMs in glioma, sensed lactate stimulation and fostered HMGB1 secretion via the cAMP/PKA/CREB signaling pathway. Blocking this feedback loop presents a promising therapeutic strategy for GBM.

Enhanced Therapeutic Potential of Hybrid Exosomes Loaded with Paclitaxel for Cancer Therapy.

The advancement of exosome studies has positioned engineered exosomes as crucial biomaterials for the development of advanced drug delivery systems. This study focuses on developing a hybrid exosome system by fusing mesenchymal stem cells (MSCs) exosomes with folate-targeted liposomes. The aim was to improve the drug loading capacity and target modification of exosome nanocarriers for delivering the first-line chemotherapy drug paclitaxel (PTX) and its effectiveness was assessed through cellular uptake studies to evaluate its ability to deliver drugs to tumor cells in vitro. Additionally, in vivo experiments were conducted using a CT26 tumor-bearing mouse model to assess the therapeutic efficacy of hybrid exosomes loaded with PTX (ELP). Cellular uptake studies demonstrated that ELP exhibited superior drug delivery capabilities to tumor cells in vitro. Moreover, in vivo experiments revealed that ELP significantly suppressed tumor growth in the CT26 tumor-bearing mouse model. Notably, for the first time, we examined the tumor microenvironment following intratumoral administration of ELP. We observed that ELP treatment activated CD4+ and CD8+ T cells, reduced the expression of M2 type tumor-associated macrophages (TAMs), polarized TAMs towards the M1 type, and decreased regulatory T cells (Tregs). Our research highlights the considerable therapeutic efficacy of ELP and its promising potential for future application in cancer therapy. The development of hybrid exosomes presents an innovative approach to enhance drug delivery and modulate the tumor microenvironment, offering exciting prospects for effective cancer treatment strategies.

Abstract 6758: Eradication of tumors and development of immunity from pHLIP-targeted intracellular delivery of STINGa

Abstract Activation of the stimulator of interferon genes (STING) pathway causes the release of factors that trigger the immune response in the tumor microenvironment (TME). Transient strong activation leads to immune activation and tumor suppression, while prolonged weak activation leads to immuno-suppression promoting tumor development. Reprogramming of M2-type tumor-associated macrophages (M2-TAMs) toward an M1 phenotype, suppression of cancer-associated fibroblasts (CAFs) and myeloid-derived suppressor cells (mMDSCs), as well as activation of dendritic cells (DCs) to train T-cells are advantageous, while the activation of STING pathway in T-cells is leads to their apoptosis. Thus, targeted delivery of STINGa to specific cells with TME to induce strong transient activation may be a key to therapy. We have developed a delivery approach that uses one or two pH Low Insertion Peptides (pHLIPs) that collaborate in the targeted intracellular delivery of a STINGa. STINGa were conjugated with pHLIPs via S-S cleavable self-immolating linkers. Biophysical studies were performed to ensure proper interactions of pHLIP-STINGa agents with membrane lipid bilayers. pHLIPs extend the lifetime of STINGa in the blood and target them to acidic cancer and stromal cells (CAFs), as well as M2-TAMs, mMDSCs and DCs within TME. The targeting results in selective cytokine activation in tumors with no systemic activation, which triggers the eradication of small (100 mm3) and large (400-700 mm3) CT26 tumors in mice after one or two doses of pHLIP-STINGa. The tumor stroma was destroyed, intratumoral hemorrhage developed, and the level of acidity within the TME was reduced. No tumors developed in mice re-challenged by an additional injection of cancer cells in a 90-day experiment. Thus, targeted delivery of STINGa to cancer cells, tumor stroma and TAMs induces activation of signaling, potentially resulting in the recruitment and infiltration of T- and NK-cells, which gain access to the tumor core. The cytotoxic activity of T- and NK-cells is not impaired by the acidic environment and immune memory is developed. The pHLIP technology may allow transformation of immuno-activating agents into more potent therapeutics, since pHLIP can target and deliver these agents to cancer cells, tumor stroma and myeloid cells. Citation Format: Michael DuPont, Anna Moshnikova, Marissa Iraca, Craig Klumpp, Hannah Visca, Dana Allababidi, Phoebe Pelzer, Donald Engelman, Oleg Andreev, Yana Reshetnyak. Eradication of tumors and development of immunity from pHLIP-targeted intracellular delivery of STINGa [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6758.

Abstract 2701: EREG reprograms M2-tumor-associated macrophage and suppresses colorectal cancer cell proliferation and migration

Abstract Colorectal cancer (CRC) represents a common malignancy within the digestive system, linked to significant morbidity and mortality rates. Tumors exert diverse effects on immune modulation, facilitating progression and metastasis. Amid the abundant immune cell populace within the tumor microenvironment (TME), tumor-associated macrophages (TAMs) play a pivotal role. M2-type TAMs actively foster tumor growth and engage in immune suppression by fostering angiogenesis and tissue restructuring. In a previous study, we analyzed single-cell RNA-sequencing (scRNA-seq) data derived from colorectal cancer patients. We observed an enrichment of the secreted phosphoprotein 1 (SPP1)+ macrophage fraction in both the tumor core and border when compared to normal tissues. Additionally, epiregulin (EREG), an epidermal growth factor receptor (EGFR) ligand, was found to be upregulated in SPP1+ macrophages, correlating with tumor growth and metastatic tendencies. Research on EREG is important because it has been found to play a significant role in cancer progression, including colorectal cancer. Understanding the mechanisms by which EREG contributes to tumor growth and metastasis can provide valuable insights into the development of targeted therapies for cancer treatment. Our hypothesis centered around the impact of EREG from TAMs on tumor progression. To identify TAM-specific EREG and its functional significance, we isolated tumor-associated macrophages from CT26 (mouse colorectal carcinoma)-bearing tumors and cultured them with conditioned and normal mediums. Subsequently, we examined EREG expression at the gene level using quantitative PCR. Our findings indicated significantly higher levels of EREG in the conditioned medium compared to the normal medium, suggesting an association between EREG and cancer cells. Remarkably, our investigation also confirmed that EREG has the capacity to reshape macrophage signatures. Notably, the surface marker CD206, characteristic of M2 macrophages, exhibited a significant decrease, while iNOS, a marker for M1 macrophages, showed an increase upon EREG depletion. We evaluated the interaction between CT26 cancer cells and TAMs using indirect co-culture methods. EREG-depleted TAMs were plated in the top chamber, while CT26 colorectal cancer cells were seeded at the bottom. Our results validated that knocking down EREG in TAMs not only hindered the proliferation of cancer cells but also affected the behavior of TAMs. Additionally, the reduced EREG expression in TAMs hindered the migration of cancer cells. These findings strongly suggest that targeting EREG could effectively restrain the growth and metastasis of colorectal cancer cells by reprogramming macrophage polarization in the TME. Consequently, this indicates its potential as a promising therapeutic target in colorectal cancer. Citation Format: Ah-Ran Yu, Seijong Kim, Yelin Jeong, Hye Kyung Hong, Yong Jae Shin, Yong Beom Cho. EREG reprograms M2-tumor-associated macrophage and suppresses colorectal cancer cell proliferation and migration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2701.

Enhancing photodynamic immunotherapy by reprograming the immunosuppressive tumor microenvironment with hypoxia relief

Tumor hypoxia impairs the generation of reactive oxygen species and the induction of immunogenic cell death (ICD) for photodynamic therapy (PDT), thus impeding its efficacy and the subsequent immunotherapy. In addition, hypoxia plays a critical role in forming immunosuppressive tumor microenvironments (TME) by regulating the infiltration of immunosuppressive tumor-associated macrophages (TAMs) and the expression of programmed death ligand 1 (PD-L1). To simultaneously tackle these issues, a MnO2-containing albumin nanoplatform co-delivering IR780, NLG919, and a paclitaxel (PTX) dimer is designed to boost photodynamic immunotherapy. The MnO2-catalyzed oxygen supply bolsters the efficacy of PDT and PTX-mediated chemotherapy, collectively amplifying the induction of ICD and the expansion of tumor-specific cytotoxic T lymphocytes (CTLs). More importantly, hypoxia releif reshapes the immunosuppressive TME via down-regulating the intratumoral infiltration of M2-type TAMs and the PD-L1 expression of tumor cells to enhance the infiltration and efficacy of CTLs in combination with immune checkpoint blockade (ICB) by NLG919, consequently eradicating primary tumors and almost completely preventing tumor relapse and metastasis. This study sets an example of enhanced immunotherapy for breast cancers through dual ICD induction and simultaneous immunosuppression modulation via both hypoxia relief and ICB, providing a strategy for the treatment of other hypoxic and immunosuppressive cancers.

Exosome-mediated communication between gastric cancer cells and macrophages: implications for tumor microenvironment.

Gastric cancer (GC) is a malignant neoplasm originating from the epithelial cells of the gastric mucosa. The pathogenesis of GC is intricately linked to the tumor microenvironment within which the cancer cells reside. Tumor-associated macrophages (TAMs) primarily differentiate from peripheral blood monocytes and can be broadly categorized into M1 and M2 subtypes. M2-type TAMs have been shown to promote tumor growth, tissue remodeling, and angiogenesis. Furthermore, they can actively suppress acquired immunity, leading to a poorer prognosis and reduced tolerance to chemotherapy. Exosomes, which contain a myriad of biologically active molecules including lipids, proteins, mRNA, and noncoding RNAs, have emerged as key mediators of communication between tumor cells and TAMs. The exchange of these molecules via exosomes can markedly influence the tumor microenvironment and consequently impact tumor progression. Recent studies have elucidated a correlation between TAMs and various clinicopathological parameters of GC, such as tumor size, differentiation, infiltration depth, lymph node metastasis, and TNM staging, highlighting the pivotal role of TAMs in GC development and metastasis. In this review, we aim to comprehensively examine the bidirectional communication between GC cells and TAMs, the implications of alterations in the tumor microenvironment on immune escape, invasion, and metastasis in GC, targeted therapeutic approaches for GC, and the efficacy of potential GC drug resistance strategies.

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.

Curcumin Suppresses M2 Macrophage-derived Paclitaxel Chemoresistance through Inhibition of PI3K-AKT/STAT3 Signaling.

Breast cancer is the leading cancer in women worldwide. The development of chemoresistance that leads to recurrence and mortality remains a major concern. M2-type tumor-associated macrophages (TAMs), present in the breast tumor microenvironment, secrete various cytokines and growth factors that induce chemoresistance. Curcumin, isolated from Curcuma longa, is known to sensitize cancer cells and increase the efficacy of standard chemotherapeutic agents. However, the effect of curcumin on the chemoresistancegenerating ability of M2 TAMs is not known. The study aimed to determine whether curcumin could modulate M2 macrophages and suppress their ability to induce resistance to paclitaxel in breast cancer cells. THP-1 cells were differentiated to M2 macrophages using PMA and IL-4/IL-13 in the presence or absence of curcumin in vitro. The effect of the conditioned medium of M2 macrophages on inducing resistance towards paclitaxel in MCF-7 or MDA-MB-231 cells was analyzed by cell proliferation assay, cell cycle analysis, wound healing and transwell migration assays. RT-PCR analysis was used to determine the mRNA expression of anti-inflammatory cytokines in M2 macrophages. The effect of curcumin on TGF-β, pAKT, and pSTAT3 in M2 macrophages was analyzed by western blotting. Our data revealed that the M2 macrophages polarized in the presence of curcumin lacked the ability to generate chemoresistance to paclitaxel in breast cancer cell lines. Transcriptomic analysis revealed the expression of TGF-β to be highest amongst M2 macrophage-secreted cytokines. We observed that purified recombinant TGF-β generated chemoresistance in breast cancer cells. We found that curcumin treatment abrogated the expression of TGF-β in M2 macrophages and suppressed their ability to induce chemoresistance in breast cancer cells. STITCH analysis showed strong interaction between curcumin and AKT/STAT3 pathway. Mechanistically, curcumin inhibited PI3K/AKT/STAT3 signaling in M2 macrophages. Western blot analysis revealed that M2 TAM CM, but not curcumin-treated macrophage CM, activated COX2/NF-κB in breast cancer cells. Our results showed that curcumin reduced the chemoresistance-generating ability of M2 TAMs. The study has revealed a non-cancer cell-autonomous mechanism by which curcumin partly overcomes the chemoresistance of paclitaxel in breast cancer.

VEGFA/NRP-1/GAPVD1 axis promotes progression and cancer stemness of triple-negative breast cancer by enhancing tumor cell-macrophage crosstalk.

Triple-negative breast cancer (TNBC) has long been considered a major clinical challenge due to its aggressive behavior and poor prognosis. Cancer stem cells (CSCs) are known as the main cells responsible for tumor origination, progression, recurrence and metastasis. Here, we report that M2-type tumor-associated macrophages (TAMs) contribute to cancer stemness in TNBC cells via the secretion of VEGFA. Reciprocally, elevated VEGFA expression by TAM-educated TNBC cells acts as a regulator of macrophage polarization, therefore constitute a feed-back loop between TNBC cells and TAMs. Mechanistically, VEGFA facilitates the CSC phenotype via the NRP-1 receptor and downstream GAPVD1/Wnt/β-catenin signaling pathway in TNBC cells. Our study underscores the crosstalk between TNBC cells and TAMs mediated by VEGFA and further clarifies the role and underlying mechanisms of the VEGFA/NRP-1/GAPVD1 axis in regulating cancer stemness. We also document an immunosuppressive function of VEGFA in the tumor microenvironment (TME). Therefore, the present study indicates crosstalk between TNBC cells and TAMs induced by VEGFA and provides a potential implication for the combination of immunotherapy and VEGFA-targeted agents in TNBC therapy.

Metformin Alters Tumor Immune Microenvironment, Improving the Outcomes of Breast Cancer Patients With Type 2 Diabetes Mellitus.

This study investigated the clinical effect of metformin on breast cancer patients with preexisting type 2 diabetes mellitus (T2DM). We analyzed 177 patients with T2DM who underwent breast cancer surgery and assessed tumor-associated macrophages (TAMs) and tumor-infiltrating lymphocytes (TILs) in patients who underwent tumor resection with or without metformin treatment using multiplex immunohistochemistry (IHC). Patients who received metformin either pre- or postoperatively exhibited reduced distant organ recurrence and improved postoperative recurrence-free survival compared to those of patients who did not. Additionally, in a subgroup of 40 patients receiving preoperative systemic therapy, metformin treatment was associated with increased rates of pathological complete response. IHC analysis revealed significantly lower levels of cluster of differentiation (CD) 68(+) CD163(+) M2-type TAMs (p < 0.01) but higher CD3(+) and CD8(+) TIL densities in the metformin-treated group compared with the same parameters in those without metformin treatment, with a significant difference in the CD8(+)/CD3(+) TIL ratio (p < 0.01). Despite the constraints posed by our small sample size, our findings suggest a potential role for metformin in modulating the immunological microenvironment, which may contribute to improved outcomes in diabetes patients with breast cancer.

C-C chemokine receptor 4 (CCR4)-positive regulatory T cells interact with tumor-associated macrophages to facilitate metastatic potential after radiation

PurposeOur previous study revealed that elevated C-C motif chemokine ligand 2 (CCL2) secretion by irradiated cancer cells recruited C-C motif chemokine receptor 2 (CCR2)-positive myeloid cells and polarized M2-type tumor-associated macrophages (TAMs), promoting lung metastasis in an established mouse model. This study investigated the impact of CCL2 and TAMs on adaptive immunity. MethodsWe assessed the influence of CCL2 and TAMs on adaptive immunity through two ectopic allograft mouse models constructed with MB49 bladder cancer cells and Lewis lung carcinoma cells. Both models exhibited delayed primary tumor growth following radiation therapy (RT), but RT promoted the development of pulmonary metastases in C57BL/6 mice. Additionally, we employed a direct coculture system to investigate the interaction between macrophages and target cells in the context of adaptive immunity. ResultsC-C motif chemokine receptor 4 (CCR4)-positive regulatory T cells (Tregs) were recruited to the postirradiated tumor microenvironment (TME). Utilizing a CCR4 antagonist to inhibit CCL2-CCR4 activation reversed the infiltration of CCR4 + Tregs and reduced the incidence of pulmonary metastases. In addition, a positive feedback loop between M2-type TAMs and Tregs was observed. The combined blockade of the CCL2-CCR4 and CCL2-CCR2 signaling pathways further decreased the risk of RT-promoted lung metastasis. ConclusionThe recruitment of CCR4 + Tregs to the postirradiated TME increases the metastatic potential of tumor cells through increased interactions with M2-type TAMs. A significant reduction in post-RT lung metastases in ectopic mouse models was achieved by disrupting the recruitment of both CCR4 + Tregs and CCR2 + myeloid cells, which are TAM precursors.

Chimeric Peptide-Engineered Self-Delivery Nanomedicine for Photodynamic-Triggered Breast Cancer Immunotherapy by Macrophage Polarization.

A systemic treatment strategy is urgently demanded to suppress the rapid growth and easy metastasis characteristics of breast cancer. In this work, a chimeric peptide-engineered self-delivery nanomedicine (designated as ChiP-CeR) for photodynamic-triggered breast cancer immunotherapy by macrophage polarization. Among these, ChiP-CeR is composed of the photosensitizer of chlorine e6 (Ce6)and the TLR7/8 agonist of lmiquimod (R837), which is further modified with tumor matrix targeting peptide (Fmoc-K(Fmoc)-PEG8-CREKA. ChiP-CeR is preferred to actively accumulate at the tumor site via specific recognition of fibronectin, which can eradicate primary tumor growth through photodynamic therapy (PDT). Meanwhile, the destruction of primary tumors would trigger immunogenic cell death (ICD) effects to release high-mobility group box-1(HMGB1)and expose calreticulin (CRT). Moreover, ChiP-CeR can also polarize M2-type tumor-associated macrophages (TAMs) into M1-type TAMs, which can activate T cell antitumor immunity in combination with ICD. Overall, ChiP-CeR possesses superior antitumor effects against primary and lung metastatic tumors, which provide an applicable nanomedicine and a feasible strategy for the systemic management of metastatic breast cancer.

Multifunctional Tumor-Targeting Carbon Dots for Tumor Microenvironment Activated Ferroptosis and Immunotherapy in Cancer Treatment.

As an emerging cancer treatment strategy, ferroptosis is distinguished by the perturbation of lipid metabolism equilibrium and the accumulation of lipid peroxidation. However, the efficacy is consistently hindered by excessive GSH in the tumor microenvironment (TME). Here, this work designed and prepared multifunctional tumor-targeting carbon dots (FG-CDs@Cu) for ferroptosis and immunotherapy. Cu2+ in FG-CDs@Cu rapidly depletes high concentrations of GSH and inhibits glutathione peroxidase 4 (GPX4) expression in an acidic TME. Meanwhile, the generated Cu+ produced reactive oxygen species (ROS) through Fenton-like reaction. Due to the high efficiency of ROS production and GSH depletion, ferroptosis mediated by oxidative stress is significantly enhanced by FG-CDs@Cu in vivo, which can induce immunogenic cell death and promote CD8+ T cell infiltration. Meanwhile, the generated O2 effectively improves the hypoxic environment of the cells and leads to the reduction of hypoxia factor-1α (HIF-1α) expression, which activates the transformation of tumor-promoting M2-type tumor-associated macrophages (TAMs) to tumor-inhibiting M1-type TAMs, further enhancing the immune response and ferroptosis. The in vivo tests suggested that FG-CDs@Cu could efficiently suppress tumor growth in the mouse model and did not cause obvious toxicity. The combination with antiprogrammed death-ligand 1 (αPD-L1) synergy immune therapy could effectively restrain the growth of distal tumors, suggesting the significant potential of FG-CDs@Cu in augmenting ferroptosis and immunotherapy for efficacious cancer treatment.

Identification and validation of necroptosis-related gene signatures to predict clinical outcomes and therapeutic responses in acute myeloid leukemia.

Necroptosis is a tightly regulated form of necrotic cell death that promotes inflammation and contributes to disease development. However, the potential roles of necroptosis-related genes (NRGs) in acute myeloid leukemia (AML) have not been elucidated fully. We conducted a study to identify a robust biomarker signature for predicting the prognosis and immunotherapy efficacy based on NRGs in AML. We analyzed the genetic and transcriptional alterations of NRGs in 151 patients with AML. Then, we identified three necroptosis clusters. Moreover, a necroptosis score was constructed and assessed based on the differentially expressed genes (DEGs) between the three necroptosis clusters. Three necroptosis clusters were correlated with clinical characteristics, prognosis, the tumor microenvironment, and infiltration of immune cells. A high necroptosis score was positively associated with a poor prognosis, immune-cell infiltration, expression of programmed cell death 1/programmed cell death ligand 1 (PD-1/PD-L1), immune score, stromal score, interferon-gamma (IFNG), merck18, T-cell dysfunction-score signatures, and cluster of differentiation-86, but negatively correlated with tumor immune dysfunction and exclusion score, myeloid-derived suppressor cells, and M2-type tumor-associated macrophages. Our observations indicated that a high necroptosis score might contribute to immune evasion. More interestingly, AML patients with a high necroptosis score may benefit from treatment based on immune checkpoint blockade. Consequently, our findings may contribute to deeper understanding of NRGs in AML, and facilitate assessment of the prognosis and treatment strategies.

Biomimetic inducer enabled dual ferroptosis of tumor and M2-type macrophages for enhanced tumor immunotherapy

Tumor-associated macrophages (TAMs) are abundant in the tumor microenvironment which promotes the formation of the immunosuppressive tumor microenvironment (ITME) through multiple mechanisms, severely counteracting the therapeutic efficacy of immunotherapy. In this study, a novel biomimetic ferroptosis inducer (D@FMN-M) capable of ITME regulation for enhanced cancer ferroptosis immunotherapy is reported. Upon tumor accumulation of D@FMN-M, the intratumoral mild acidity triggers the biodegradation of Fe-enriched nanocarriers and the concurrent co-releases of dihydroartemisinin (DHA) and Fe3+. The released Fe3+ is reduced to Fe2+ by consuming intratumoral glutathione (GSH), which promotes abundant free radical generation via triggering Fenton and Fe2+-DHA reactions, thus inducing ferroptosis of both cancer cells and M2-type TAMs. Resultantly, the anticancer immune response is strongly activated by the massive tumor-associated antigens released by ferroptositic cancer cells. Also importantly, the ferroptosis-sensitive M2-type TAMs will be either damaged or gradually domesticated to ferroptosis-resistant M1 TAMs under the ferroptosis stress, favoring the normalization of ITME and finally amplifying cancer ferroptosis immunotherapeutic efficacy. This work provides a novel strategy for ferroptosis immunotherapy of solid tumors featuring TAMs infiltration and immunosuppression by inducing dual ferroptosis of tumor cells and M2-type TAMs.

Tumor cell SPTBN1 inhibits M2 polarization of macrophages by suppressing CXCL1 expression.

Tumor-associated macrophages (TAMs) are the most abundant immune cells in the tumor microenvironment, and the M2-type TAMs can promote tumor growth, invasion and angiogenesis, and suppress antitumor immune responses. It has been reported that spectrin beta, non-erythrocytic 1 (SPTBN1) may inhibit the infiltration of macrophages in Sptbn1+/- mouse liver, but whether tumor SPTBN1 affects TAMs polarization remains unclear. This study investigated the effect and mechanism of tumor cell SPTBN1 on polarization and migration of TAMs in hepatoma and breast cancer. By analyzing tumor immune databases, we found a negative correlation between SPTBN1 and abundance of macrophages and myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment. Byreverse transcription-quantitative real-time PCRassays and cell migration assays, the migration and M2 polarization of macrophages were enhanced by the culture medium from hepatocellular carcinoma cell line PLC/PRF/5, SNU449, and breast cancer cell line MDA-MB-231 with SPTBN1 suppression, which could be reversed by CXCL1 neutralizing antibody MAB275. Meanwhile, the ability of migration and colony formation of PLC/PRF/5, SNU449, and MDA-MB-231 cells were promoted when coculture with M2 macrophages. We also found that SPTBN1 regulated CXCL1 through p65 by cytoplasmic-nuclear protein isolation experiments and ChIP-qPCR. Our data suggest that tumor cell SPTBN1 inhibits migration and M2-type polarization of TAMs by reducing the expression and secretion of CXCL1 via inhibiting p65 nuclear localization.

Dauricine regulates prostate cancer progression by inhibiting PI3K/AKT-dependent M2 polarization of macrophages

M2 type tumor-associated macrophages, an essential component of the tumor microenvironment (TME), have been proved to contribute to tumor metastasis. Dauricine (Dau) has recently received widespread attention due to its multiple targets and low price. However, the effect of Dau on macrophage polarization of TME remains unclear. In this study, we investigated the effect of Dau on prostate cancer (PCa) metastasis and specifically its correlation to macrophage polarization. Our results showed that Dau efficiently suppressed M2 polarization of macrophages induced by interleukin (IL) −4 and IL-13. Mechanistically, Dau inhibited the activity of PI3K/AKT signaling pathway, which subsequently suppressed macrophage differentiation to M2 type. Importantly, our study indicated that Dau decreased the release of chitinase 3-like protein 1 (CHI3L1) from M2 macrophages, which ultimately inhibited the M2 macrophage-mediated progression of PCa cells in vitro and in vivo. Taken together, our data demonstrated that Dau suppressed M2 polarization of macrophages via downregulation of the PI3K/AKT signaling pathway, in turn, preventing proliferation, epithelial–mesenchymal transition, migration, and invasion of PCa cells. Thus, this study reveals a previously unrecognized function of Dau in inhibition of PCa progression via intervention in M2 polarization of macrophages.