Unconventional p65/p52 NF-κB module regulates key tumor microenvironment-related genes in breast tumor-associated macrophages (TAMs).

The complex heterogeneity of tumor microenvironment (TME) of triple-negative breast cancer (TNBC) presents a significant obstacle to cytotoxic immune response and successful treatment, building up one of the most hostile oncological phenotypes. Among the most abundant TME components, tumor-associated macrophages (TAMs) have pivotal pro-tumoral functions, involving discordant roles for the nuclear factor kappa-B (NF-κB) transcription factors and directing to higher levels of pathway complexity. In both resting macrophages and TAMs, we recently revealed the existence of the uncharacterized NF-κB p65/p52 dimer. In the present study, we demonstrated its enhanced active nuclear localization in TAMs and validated selected immune target genes as directly regulated by dimer binding on DNA sequences. We demonstrated by ChIP-qPCR that p65/p52 enrichment on HSPG2 and CSF-1 regulatory regions is strictly dependent on macrophage polarization and tumor environment. Our data provide novel mechanisms of transcriptional regulation in TAMs, orchestrated by the varied and dynamic nature of NF-κB combinations, which needs to be considered when targeting this pathway in cancer therapies. Our results offer p65/p52, together with identified regulatory regions on genes impacting macrophage behavior and tumor biology, as novel molecular targets for TNBC, aimed at modulating TAMs functions towards anti-tumoral phenotypes and thus improving cancer treatment outcomes.

Drug resistance and tumor immune microenvironment: An overview of current understandings (Review).

The use of antitumor drugs represents a reliable strategy for cancer therapy. Unfortunately, drug resistance has become increasingly common and contributes to tumor metastasis and local recurrence. The tumor immune microenvironment (TME) consists of immune cells, cytokines and immunomodulators, and collectively they influence the response to treatment. Epigenetic changes including DNA methylation and histone modification, as well as increased drug exportation have been reported to contribute to the development of drug resistance in cancers. In the past few years, the majority of studies on tumors have only focused on the development and progression of a tumor from a mechanistic standpoint; few studies have examined whether the changes in the TME can also affect tumor growth and drug resistance. Recently, emerging evidence have raised more concerns regarding the role of TME in the development of drug resistance. In the present review, it was discussed how the suppressive TME adapts to drug resistance characterized by the cooperation of immune cells, cytokines, immunomodulators, stromal cells and extracellular matrix. Furthermore, it was reviewed how these immunological or metabolic changes alter immuno­surveillance and thus facilitate tumor drug resistance. In addition, potential targets present in the TME for developing novel therapeutic strategies to improve individualized therapy for cancer treatment were revealed.

Tumor­associated macrophages activated in the tumor environment of hepatocellular carcinoma: Characterization and treatment (Review).

Hepatocellular carcinoma (HCC) tissue is rich in dendritic cells, T cells, B cells, macrophages, natural killer cells and cellular stroma. Together they form the tumor microenvironment (TME), which is also rich in numerous cytokines. Tumor­associated macrophages (TAMs) are involved in the regulation of tumor development. TAMs in HCC receive stimuli in different directions, polarize in different directions and release different cytokines to regulate the development of HCC. TAMs are mostly divided into two cell phenotypes: M1 and M2. M1 TAMs secrete pro­inflammatory mediators, and M2 TAMs secrete a variety of anti­inflammatory and pro­tumorigenic substances. The TAM polarization in HCC tumors is M2. Both direct and indirect methods for TAMs to regulate the development of HCC are discussed. TAMs indirectly support HCC development by promoting peripheral angiogenesis and regulating the immune microenvironment of the TME. In terms of the direct regulation between TAMs and HCC cells, the present review mainly focuses on the molecular mechanism. TAMs are involved in both the proliferation and apoptosis of HCC cells to regulate the quantitative changes of HCC, and stimulate the related invasive migratory ability and cell stemness of HCC cells. The present review aims to identify immunotherapeutic options based on the mechanisms of TAMs in the TME of HCC.

Inhibition of STAT3 by 2-Methoxyestradiol suppresses M2 polarization and protumoral functions of macrophages in breast cancer.

BACKGROUND: Breast cancer metastasis remains the leading cause of cancer-related deaths in women worldwide. Infiltration of tumor-associated macrophages (TAMs) in the tumor stroma is known to be correlated with reduced overall survival. The inhibitors of TAMs are sought after for reprogramming the tumor microenvironment. Signal transducer and activator of transcription 3 (STAT3) is well known to contribute in pro-tumoral properties of TAMs. 2-Methoxyestradiol (2ME2), a potent anticancer and antiangiogenic agent, has been in clinical trials for treatment of breast cancer. Here, we investigated the potential of 2ME2 in modulating the pro-tumoral effects of TAMs in breast cancer. METHODS: THP-1-derived macrophages were polarized to macrophages with or without 2ME2. The effect of 2ME2 on macrophage surface markers and anti-inflammatory genes was determined by Western blotting, flow cytometry, immunofluorescence, qRT‒PCR. The concentration of cytokines secreted by cells was monitored by ELISA. The effect of M2 macrophages on malignant properties of breast cancer cells was determined using colony formation, wound healing, transwell, and gelatin zymography assays. An orthotopic model of breast cancer was used to determine the effect of 2ME2 on macrophage polarization and metastasis in vivo. RESULTS: First, our study found that polarization of monocytes to alternatively activated M2 macrophages is associated with the reorganization of the microtubule cytoskeleton. At lower concentrations, 2ME2 treatment depolymerized microtubules and reduced the expression of CD206 and CD163, suggesting that it inhibits the polarization of macrophages to M2 phenotype. However, the M1 polarization was not significantly affected at these concentrations. Importantly, 2ME2 inhibited the expression of several anti-inflammatory cytokines and growth factors, including CCL18, TGF-ß, IL-10, FNT, arginase, CXCL12, MMP9, and VEGF-A, and hindered the metastasis-promoting effects of M2 macrophages. Concurrently, 2ME2 treatment reduced the expression of CD163 in tumors and inhibited lung metastasis in the orthotopic breast cancer model. Mechanistically, 2ME2 treatment reduced the phosphorylation and nuclear translocation of STAT3, an effect which was abrogated by colivelin. CONCLUSIONS: Our study presents novel findings on mechanism of 2ME2 from the perspective of its effects on the polarization of the TAMs via the STAT3 signaling in breast cancer. Altogether, the data supports further clinical investigation of 2ME2 and its derivatives as therapeutic agents to modulate the tumor microenvironment and immune response in breast carcinoma.

Targeted nanomedicine for reprogramming the tumor innate immune system: From bench to bedside.

Tumor-associated innate immune cells such as tumor-associated macrophages, neutrophils, dendritic cells play a crucial role in tumor progression, angiogenesis and metastasis. These cells also control the efficacy of chemotherapy and immunotherapy by inducing drug resistance and immunosuppression, leading to therapeutic failures. Therefore, targeting the tumor-associated innate immune cells has gained high attention for the development of effective cancer therapy. Nanomedicine based strategies to target these cells are highly relevant and can be used to reprogram these cells. In this review, we discuss the fundamental roles of the tumor-associated innate immune cells in the tumor microenvironment and different strategies to modulate them. Then, nanomedicine-based strategies to target different tumor innate immune cells are explained in detail. While the clinical development of the targeted nanomedicine remains a great challenge in practice, we have provided our perspectives on various factors such as pharmaceutical aspects, preclinical testing and biological aspects which are crucial to consider before translating these targeting strategies to clinics.

Correlation of Immunomodulatory Cytokines with Tumor Volume and Cerebrospinal Fluid in Vestibular Schwannoma Patients.

Sporadic vestibular schwannomas (VSs) often exhibit slow or negligible growth. Nevertheless, some VSs increase significantly in volume within a few months or grow continuously. Recent evidence indicates a role of inflammation in promoting VS growth. Therefore, our study aimed to identify cytokines, which are associated with larger VSs. The expression of different cytokines in VS tumor samples and VS primary cultures was investigated. Additionally, the concentration of cytokines in cell culture supernatants of VS primary cultures and cerebrospinal fluid (CSF) of VS patients and healthy controls were determined. Correlation analysis of cytokine levels with tumor volume, growth rate, Koos grade, age, and hearing was examined with Spearman's-rank test. The mRNA expression of CC-chemokine ligand (CCL) 18, growth differentiation factor (GDF) 15, and interferon regulatory factor 4 correlated positively with tumor volume. Moreover, the amount of GDF15 in the cell culture supernatant of primary cells correlated positively with tumor volume. The concentrations of the cytokines CCL2, CCL5, and CCL18 and transforming growth factor beta (TGFB) 1 in the CSF of the patients were significantly different from those in the CSF controls. Inhibition of immune cell infiltration could be a putative approach to prevent and control VS growth.

The Importance of Suppressing Pathological Periostin Splicing Variants with Exon 17 in Both Stroma and Cancer.

BACKGROUND: Periostin (POSTN) is a type of matrix protein that functions by binding to other matrix proteins, cell surface receptors, or other molecules, such as cytokines and proteases. POSTN has four major splicing variants (PN1-4), which are primarily expressed in fibroblasts and cancer. We have reported that we should inhibit pathological POSTN (PN1-3), but not physiological POSTN (PN4). In particular, pathological POSTN with exon 17 is present in both stroma and cancer, but it is unclear whether the stroma or cancer pathological POSTN should be suppressed. METHODS AND RESULTS: We transplanted 4T1 cells (breast cancer) secreting POSTN with exon 17 into 17KO mice lacking POSTN exon 17 to suppress stromal POSTN with exon 17. The results show that 17KO mice had smaller primary tumors and fewer metastases. Furthermore, to suppress cancer POSTN with exon 17, 4T1 cells transfected with POSTN exon 17 skipping oligo or control oligo were transplanted from the tail vein into the lungs. The results show that POSTN exon 17 skipping oligo significantly suppressed lung metastasis. CONCLUSIONS: These findings suggest that it is important to suppress POSTN exon 17 in both stroma and cancer. Antibody targeting POSTN exon 17 may be a therapeutic candidate for breast cancer.

FAM109B plays a tumorigenic role in low-grade gliomas and is associated with tumor-associated macrophages (TAMs).

BACKGROUND: Family with sequence similarity 109, member B (FAM109B) is involved in endocytic transport and affects genetic variation in brain methylation. It is one of the important genes related to immune cell-associated diseases. In the tumor immune system, methylation can regulate tumor immunity and influence the maturation and functional response of immune cells. Whether FAM109B is involved in tumor progression and its correlation with the tumor immune microenvironment has not yet been disclosed. METHODS: A comprehensive pan-cancer analysis of FAM109B expression, prognosis, immunity, and TMB was conducted. The expression, clinical features, and prognostic value of FAM109B in low-grade gliomas (LGG) were evaluated using TCGA, CGGA, and Gravendeel databases. The expression of FAM109B was validated by qRT-PCR, immunohistochemistry (IHC), and Western blotting (WB). The relationship between FAM109B and methylation, Copy Number Variation (CNV), prognosis, immune checkpoints (ICs), and common chemotherapy drug sensitivity in LGG was explored through Cox regression, Kaplan-Meier curves, and Spearman correlation analysis. FAM109B levels and their distribution were studied using the TIMER database and single-cell analysis. The potential role of FAM109B in gliomas was further investigated through in vitro and in vivo experiments. RESULTS: FAM109B was significantly elevated in various tumor types and was associated with poor prognosis. Its expression was related to aggressive progression and poor prognosis in low-grade glioma patients, serving as an independent prognostic marker for LGG. Glioma grade was negatively correlated with FAM109B DNA promoter methylation. Immune infiltration and single-cell analysis showed significant expression of FAM109B in tumor-associated macrophages (TAMs). The expression of FAM109B was closely related to gene mutations, immune checkpoints (ICs), and chemotherapy drugs in LGG. In vitro studies showed increased FAM109B expression in LGG, closely related to cell proliferation. In vivo studies showed that mice in the sh-FAM109B group had slower tumor growth, slower weight loss, and longer survival times. CONCLUSIONS: FAM109B, as a novel prognostic biomarker for low-grade gliomas, exhibits specific overexpression in TAMs and may be a potential therapeutic target for LGG patients.

ICAT-Mediated Crosstalk Between Cervical Cancer Cells and Macrophages Promotes M2-Like Macrophage Polarization to Reinforce Tumor Malignant Behaviors.

Inhibitor of ß-catenin and T-cell factor (ICAT) is a classical inhibitor of the Wnt signaling pathway. Nonetheless, our previous work found that ICAT is overexpressed in cervical cancer (CC), resulting in the augmentation of migration and invasion capabilities of CC cells. It remains unclear what molecular mechanism underlies this phenomenon. The interaction between cancer cells and the tumor microenvironment (TME) promotes the outgrowth and metastasis of tumors. Tumor-associated macrophages (TAMs) are a major constituent of the TME and have a significant impact on the advancement of CC. Consequently, our inquiry pertains to the potential of ICAT to facilitate tumor development through its modulation of the cervical TME. In this study, we first verified that ICAT regulated the secretion of cytokines interleukin-10 (IL-10) and transforming growth factor-ß (TGF-ß) in CC cells, leading to M2-like macrophage polarization and enhancement of the migration and invasion of CC cells. Furthermore, the system of co-culturing human umbilical vein endothelial cells (HUVECs) with macrophages revealed that depending on the CC cells' overexpression or inhibition of ICAT, the vascular tube formation by HUVECs can be either increased or decreased. Overall, our study indicates that ICAT stimulates M2-like polarization of TAMs via upregulating IL-10 and TGF-ß, which results in increased neovascularization, tumor metastasis, and immunosuppression in CC. In upcoming times, inhibiting crosstalk between CC cells and TAMs may be a possible strategy for CC therapy.

Melatonin downregulates angiogenesis and lymphangiogenesis by regulating tumor-associated macrophages via NLRP3 inflammasomes in lung adenocarcinoma.

Tumor-associated macrophages (TAMs), present within the tumor microenvironment (TME), strictly modulate tumor angiogenesis and lymphangiogenesis. Nevertheless, the associated signaling networks and candidate drug targets for these events remains to be elucidated. Given its antioxidative activities, we speculated that melatonin may reduce pyroptosis, and thereby modulate both angiogenesis and lymphangiogenesis. We revealed that a co-culture of A549 cells and THP-1 macrophages strongly enhanced expressions of the NLRP3 inflammasome axis members, and augmented angiogenesis and lymphangiogenesis. Next, we overexpressed NLRP3 in the A549 cells, and demonstrated that excess NLRP3 expression substantially upregulated VEGF and CXCL cytokine expressions, and enhanced lymphatic endothelial cells (LECs) tube formation. In contrast, NLRP3 inhibition produced the opposite effect. In addition, relative to controls, melatonin administration strongly inhibited the NLRP3 inflammasome axis, as well as angiogenesis and lymphangiogenesis in the co-culture system. Subsequent animal experiments using a Lewis Lung Carcinoma (LLC) subcutaneous tumor model in mice corroborate these findings. Melatonin treatment and NLRP3 knockdown significantly inhibit tumor growth and downregulate NLRP3 and IL-1ß expression in tumor tissues. Furthermore, melatonin downregulates the expression of angiogenic and lymphangiogenic markers in tumor tissues. Taken together, the evidence suggested that a THP-1 macrophage and A549 cell co-culture stimulates angiogenesis and lymphangiogenesis via the NLRP3 axis. Melatonin protected against the TAMs- and NLRP3 axis-associated promotion of the aforementioned events in vitro and in vivo. Hence, melatonin is a promising candidate for managing for tumor-related angiogenesis and lymphangiogenesis in lung adenocarcinoma.

Proteomic profiling of oleamide-mediated polarization in a primary human monocyte-derived tumor-associated macrophages (TAMs) model: a functional analysis.

Background: Tumor-associated macrophages (TAMs) play a critical function in the development of tumors and are associated with protumor M2 phenotypes. Shifting TAMs towards antitumor M1 phenotypes holds promise for tumor immunotherapy. Oleamide, a primary fatty acid amide, has emerged as a potent anticancer and immunomodulatory compound. However, the regulatory effects of oleamide on TAM phenotypes remain unclear. Methods: We used real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) techniques to study the influence of oleamide on primary human monocyte-derived TAM phenotypes, and we investigated the protein expression profiles based on mass spectrometry to analyze the effect of oleamide on macrophage polarization. Moreover, the advantageous binding scores between oleamide and these target candidate proteins are examined using molecular docking. Results: Our study revealed that oleamide effectively suppressed the M2-like TAM phenotype by reducing interleukin (IL)-10 production and downregulating M2-like markers, including vascular endothelial growth factor A (VEGFA), MYC proto-oncogene, bHLH transcription factor (c-Myc), and mannose receptor C-type 1 (CD206). Moreover, the conditioned medium derived from oleamide-treated TAMs induces apoptosis of MDA-MB-231 breast cancer cells. Proteomic analysis identified 20 candidate up- and down-regulation proteins targeted by oleamide, showing modulation activity associated with the promotion of the M1-like phenotype. Furthermore, molecular docking demonstrated favorable binding scores between oleamide and these candidate proteins. Collectively, our findings suggest that oleamide exerts a potent antitumor effect by promoting the antitumor M1-like TAM phenotype. These novel insights provide valuable resources for further investigations into oleamide and macrophage polarization which inhibit the progression of breast cancer, which may provide insight into immunotherapeutic approaches for cancer.

Targeting tumor-associated macrophages with nanocarrier-based treatment for breast cancer: A step toward developing innovative anti-cancer therapeutics.

Tumor-associated macrophages (TAMs) promote tumor advancement in many ways, such as inducing angiogenesis and the formation of new blood vessels that provide tumors with nourishment and oxygen. TAMs also facilitate tumor invasion and metastasis by secreting enzymes that degrade the extracellular matrix and generating pro-inflammatory cytokines that enhance the migration of tumor cells. TAMs also have a role in inhibiting the immune response against malignancies. To accomplish this, they release immunosuppressive cytokines such as IL-10, and TAMs can hinder the function of T cells and natural killer cells, which play crucial roles in the immune system's ability to combat cancer. The role of TAMs in breast cancer advancement is a complex and dynamic field of research. Therefore, TAMs are a highly favorable focus for innovative breast cancer treatments. This review presents an extensive overview of the correlation between TAMs and breast cancer development as well as its role in the tumor microenvironment (TME) shedding light on their impact on tumor advancement and immune evasion mechanisms. Notably, our study provides an innovative approach to employing nanomedicine approaches for targeted TAM therapy in breast cancer, providing an in-depth overview of recent advances in this emerging field.

Establishment of tumor microenvironment following bisphenol A exposure in the testis.

Although detrimental roles of bisphenol A (BPA) in xenoestrogen target organs, testis and epididymis, and male fertility are well-documented, disruption of the immune privilege system in the male reproductive tract following BPA exposure remains poorly understood. Therefore, this study aimed to explore the precise mechanisms of BPA in interfering immune privilege in the testis on RNA sequencing results. CD-1 male mice were daily treated no-observed-adverse-effect (NOAEL, 5 mg BPA/kg BW) and lowest-observed-adverse-effects (LOAEL, 50 mg BPA/kg BW) of BPA by oral gavage for 6 weeks. Following the LOAEL exposure, the expression of immune response-associated transcripts was upregulated in the testis. Moreover, BPA switch the testicular microenvironment to tumor friendly through the recruitment of tumor associated macrophages (TAMs), which can produce both anti- and pro-inflammatory cytokines, such as TNF-α, TLR2, IL-10, and CXCL9. Number of testicular blood vessels were approximately 2-times increased by upregulation of matrix metallopeptidase 2 in TAMs and upregulation of AR expression in the nucleus of Leydig cells. Moreover, we found that the tumor-supportive environment can also be generated even though NOAEL BPA concentration due to the individual's variability in cancer susceptibility.

TBK1-Zyxin signaling controls tumor-associated macrophage recruitment to mitigate antitumor immunity.

Mechanical control is fundamental for cellular localization within a tissue, including for tumor-associated macrophages (TAMs). While the innate immune sensing pathways cGAS-STING and RLR-MAVS impact the pathogenesis and therapeutics of malignant diseases, their effects on cell residency and motility remain incompletely understood. Here, we uncovered that TBK1 kinase, activated by cGAS-STING or RLR-MAVS signaling in macrophages, directly phosphorylates and mobilizes Zyxin, a key regulator of actin dynamics. Under pathological conditions and in STING or MAVS signalosomes, TBK1-mediated Zyxin phosphorylation at S143 facilitates rapid recruitment of phospho-Zyxin to focal adhesions, leading to subsequent F-actin reorganization and reduced macrophage migration. Intratumoral STING-TBK1-Zyxin signaling was evident in TAMs and critical in antitumor immunity. Furthermore, myeloid-specific or global disruption of this signaling decreased the population of CD11b+ F4/80+ TAMs and promoted PD-1-mediated antitumor immunotherapy. Thus, our findings identify a new biological function of innate immune sensing pathways by regulating macrophage tissue localization, thus providing insights into context-dependent mitigation of antitumor immunity.

Ultrasound-mediated nanobubbles loaded with STAT6 siRNA inhibit TGF-ß1-EMT axis in LUSC cells via overcoming the polarization of M2-TAMs.

M2-like tumor-associated macrophages (M2-TAMs) are closely correlated with metastasis and poor clinical outcomes in lung squamous cell carcinoma (LUSC). Previous studies have demonstrated that STAT6 is an important signaling molecule involved in the polarization of M2-TAMs, EMT is the main way for TAMs to promote tumor progression. However, little attention has been paid to the effect of STAT6 inhibition on LUSC, and it is difficult to achieve an ideal gene silencing effect in immune cells using traditional gene transfection methods. Here, we investigated the optimal concentration of 12-myristic 13-acetate (PMA), lipopolysaccharide (LPS) for the induction of THP-1 into M1-TAMs and M2-TAMs. The expression of pSTAT6 and STAT6 was confirmed in three types of macrophages, and it was demonstrated that pSTAT6 can be used as a specific target of M2-TAMs derived from THP-1. Ultrasound-mediated nanobubble destruction (UMND) is a non-invasive and safe gene delivery technology. We also synthesized PLGA-PEI nanobubbles (NBs) to load and deliver STAT6 small interfering RNA (siRNA) into M2-TAMs via UMND. The results show that the NBs could effectively load with siRNA and had good biocompatibility. We found that UMND enhanced the transfection efficiency of siRNA, as well as the silencing effect of pSTAT6 and the inhibition of M2-TAMs. Simultaneously, when STAT6 siRNA entered M2-TAMs by UMND, proliferation, migration, invasion and EMT in LUSC cells could be inhibited via the transforming growth factor-ß1 (TGF-ß1) pathway. Therefore, our results confirm that UMND is an ideal siRNA delivery strategy, revealing its potential to inhibit M2-TAMs polarization and ultimately treat LUSC.

Co-delivery of Liposomal Ketoconazole and Bevacizumab for Synergistical Inhibition of Angiogenesis Against Endometrial Cancer.

In this study, we designed a novel formulation based on liposomes for the co-delivery of cancer-derived exosome inhibitor (ketoconazole, Keto) and angiogenesis inhibitor (bevacizumab, mAb). The designed Combo-Lipo formulation was systematically characterized, exhibiting a uniform average particle size of 100 nm, as well as excellent serum and long-term physical stabilities. The cell viability assay revealed that Combo-Lipo treatment significantly reduced the viability of cancer cells compared to free drugs. Moreover, liposomes effectively inhibited angiogenic mediators and reduced tumor immune suppressive factors. The Combo-Lipo formulation demonstrated potent downregulation of angiogenic factors and synergistic effects in suppressing their production. Furthermore, liposomes inhibited tumor-associated macrophages (TAMs), leading to decreased expression of tumor-promoting factors. Together, these findings highlighted the promising characteristics of Combo-Lipo as a therapeutic formulation, including optimal particle size, serum stability, and potent anti-cancer effects, as well as inhibition of angiogenic mediators and TAMs toward treating endometrial cancer.

Comprehensive scRNA-seq analysis to identify new markers of M2 macrophages for predicting the prognosis of prostate cancer.

BACKGROUND: Prostate cancer (PCa) has become the highest incidence of malignant tumor among men in the world. Tumor microenvironment (TME) is necessary for tumor growth. M2 macrophages play an important role in many solid tumors. This research aimed at the role of M2 macrophages' prognosis value in PCa. METHODS: Single-cell RNA-seq (scRNA-seq) data and mRNA expression data were obtained from the Gene Expression Omnibus database (GEO) and The Cancer Genome Atlas (TCGA). Quality control, normalization, reduction, clustering, and cell annotation of scRNA-seq data were preformed using the Seruat package. The sub-populations of the tumor-associated macrophages (TAMs) were analysis and the marker genes of M2 macrophage were selected. Differentially expressed genes (DEGs) in PCa were identified using limma and the immune infiltration was detected using CIBERSORTx. Then, a weighted correlation network analysis (WGCNA) was constructed to identify the M2 macrophage-related modules and genes. Integration of the marker genes of M2 macrophage from scRNA-seq data analysis and hub genes from WGCNA to select the prognostic gene signature based on Univariate and LASSO regression analysis. The risk score was calculated, and the DEGs, biological function, immune characteristics related to risk score were explored. And a predictive nomogram was constructed. CCK8, Transwell, and wound healing were used to verify cell phenotype changes after co-cultured. RESULTS: A total of 2431 marker genes of M2 macrophage and 650 hub M2 macrophage-related genes were selected based on scRNA-seq data and WGCNA. Then, 113 M2 macrophage-related genes were obtained by overlapping the scRNA-seq data and WGCNA results. Nine M2 macrophage-related genes (SMOC2, PLPP1, HES1, STMN1, GPR160, ABCG1, MAZ, MYC, and EPCAM) were screened as prognostic gene signatures. M2 risk score was calculated, the DEGs, Immune score, stromal score, ESTIMATE score, tumor purity, and immune cell infiltration, immune checkpoint expression, and responses of immunotherapy and chemotherapy were identified. And a predictive nomogram was constructed. CCK8, Transwell invasion, and wound healing further verified that M2 macrophages promoted the proliferation, invasion, and migration of PCa (p < 0.05). CONCLUSIONS: We uncovered that M2 macrophages and relevant genes played key roles in promoting the occurrence, development, and metastases of PCa and played as convincing predictors in PCa.

Construction and drug screening of Co-culture system using extrahepatic cholangiocarcinoma organoids and tumor-associated macrophages.

Patient-derived organoids (PDOs) have been proposed as a novel in vitro tumor model that can be applied to tumor research and drug screening. However, current tumor organoid models lack components of the tumor microenvironment, particularly tumor-associated macrophages(TAMs).We collected peripheral blood and tumor samples from 6 patients with extrahepatic cholangiocarcinoma(eCCA). Monocytes were induced into TAMs by cytokine and conditioned medium, and then co-cultured with tumor organoids. Our comprehensive analysis and comparison of histopathology and genomics results confirmed that this co-culture model can better capture intra- and inter-tumor heterogeneity retain the specific mutations of the original tumor. Drug sensitivity data in vitro revealed that gemcitabine and cisplatin are effective drugs for cholangiocarcinoma, but TAMs in the tumor microenvironment promote organoids growth and chemotherapy resistance. In conclusion, our organoid model of cholangiocarcinoma co-cultured with TAMs can not only shorten the model construction cycle, but also preserve the heterogeneity of original tumors to improve the accuracy of drug screening, and can also be applied to the researches of TAMs and tumors.

Magnetic resonance imaging-based radiomics for predicting infiltration levels of CD68+ tumor-associated macrophages in glioblastomas.

PURPOSE: Tumor-associated macrophages (TAMs) are important biomarkers of tumor invasion and prognosis in patients with glioblastoma. We combined the imaging and radiomics features of preoperative MRI to predict CD68+ macrophage infiltration. METHODS: Clinical, MRI image, and pathology data of 188 patients with glioblastoma were analyzed. Overall, 143 patients were included in the training (n = 101) and validation (n = 42) sets, whereas 45 patients were included in an independent test set. The optimal cut-off value (14.8%) was based on the minimum p-value formed by the Kaplan-Meier survival analysis and log-rank tests which divided patients into groups with high CD68+ TAMs (≥ 14.8%) and low CD68+ TAMs (< 14.8%). Regions of interest and radiomics features extraction were based on contrast-enhanced T1-weighted images (CE-T1WI) and T2WI. Multi-parameter stepwise regression was used to create the clinical, radiomics, and combined models, each evaluated using the receiver operating characteristic curve. Decision curve analysis was used to assess the clinical applicability of the nomogram. RESULTS: A clinical model based on the minimum apparent diffusion coefficient (ADCmin) revealed an area under the curve (AUC) of 0.768, 0.764, and 0.624 for the training set, validation set, and test set, respectively. The 2D radiomics model, based on two features, revealed an AUC of 0.783, 0.724, and 0.789 for the training, validation, and test sets, respectively. The 3D radiomics model, based on three features, revealed AUCs of 0.823, 0.811, and 0.787 for the training, validation, and test sets, respectively. The combined model, with ADCmin and radiomics features, showed the best performance, with AUCs of 0.865, 0.822, and 0.776 for the training, validation, and test sets, respectively. The calibration curve of the combined model nomogram showed good agreement between the estimated and actual probabilities. CONCLUSION: The combined model constructed using ADCmin, a quantitative imaging parameter, combined with five key radiomics features can be used to evaluate the extent of CD68+ macrophages before surgery.

TET3 is expressed in prostate cancer tumor-associated macrophages and is associated with anti-androgen resistance.

PURPOSE: The aim of this study is to investigate the expression of TET3 in prostate cancer and its effect on the efficacy of anti-androgen therapy (ADT). METHODS: The expression of TET3 in 1965 cases of prostate cancer and 493 cases of normal prostate tissues were analyzed. The CIBERSORT algorithm evaluated the abundance of 22 tumor-infiltrating immune cells in 497 prostate cancers. Subsequently, the expression of TET3 in prostate cancer TAMs was analyzed using 21,292 cells from single-cell RNA sequencing (scRNAseq). In addition, the trajectory of the differentiation process was reconstructed based on pseudotime analysis. Sensitivity prediction of prostate cancers to ADT was evaluated based on GDSC2 and CTRP databases. Another dataset GSE111177 was employed for further analysis. RESULTS: TET3 was over-expressed in prostate cancer, and the expression of TET3 in metastatic prostate cancer was higher than that in non-metastatic prostate cancer. The scRNAseq analysis of prostate cancer showed that TET3 was mainly expressed in TAM. TET3 expressed in early and active TAMs, with the activation of signaling pathways such as energy metabolism, cell communication, and cytokine production. Prostate cancer in TET3 high expression group was more sensitive to ADT drugs such as Bicalutamide and AZD3514, and was also more sensitive to chemotherapy drugs such as Cyclophosphamide, Paclitaxel, and Vincristine, and MAPK pathway inhibitors of Docetaxel and Dabrafenib. CONCLUSIONS: The efficacy of ADT in prostate cancer is related to the expression of TET3 in TAMs, and TET3 may be a potential therapeutic target for coordinating ADT.