Reprogramming Of Macrophages Research Articles

Transcriptional profiling of macrophages in situ in metastatic melanoma reveals localization-dependent phenotypes and function

SummaryModulation of immune function at the tumor site could improve patient outcomes. Here, we analyze patient samples of metastatic melanoma, a tumor responsive to T cell-based therapies, and find that tumor-infiltrating T cells are primarily juxtaposed to CD14+ monocytes/macrophages rather than melanoma cells. Using immunofluorescence-guided laser capture microdissection, we analyze transcriptomes of CD3+ T cells, CD14 + monocytes/macrophages, and melanoma cells in non-dissociated tissue. Stromal CD14+ cells display a specific transcriptional signature distinct from CD14+ cells within tumor nests. This signature contains LY75, a gene linked with antigen capture and regulation of tolerance and immunity in dendritic cells (DCs). When applied to TCGA cohorts, this gene set can distinguish patients with significantly prolonged survival in metastatic cutaneous melanoma and other cancers. Thus, the stromal CD14+ cell signature represents a candidate biomarker and suggests that reprogramming of stromal macrophages to acquire DC function may offer a therapeutic opportunity for metastatic cancers.

KLF7 Alleviates Atherosclerotic Lesions and Inhibits Glucose Metabolic Reprogramming in Macrophages by Regulating HDAC4/miR-148b-3p/NCOR1

Objectives: Atherosclerosis (AS) remains a major contributor to death worldwide. This study sought to explore the role of Krüppel-like factor 7 (KLF7) in AS lesions via regulating glucose metabolic reprogramming (GMR) in macrophages. Methods: AS mouse and cell models were established via high-fat-diet feeding and oxidized low-density lipoprotein (ox-LDL) induction. KLF7, histone deacetylase 4 (HDAC4), miR-148b-3p, and nuclear receptor corepressor 1 (NCOR1) expressions in aortic tissue and cells were detected via reverse transcription quantitative polymerase chain reaction or Western blotting. Parameters of AS lesions and mouse metabolism were detected via hematoxylin-eosin, oil red O, and Masson staining, assay kits, glucose tolerance test, and enzymatic analysis. Peritoneal macrophages of mice were isolated and cellular metabolism was detected via Seahorse metabolic flux analysis, assay kits, ELISA, and Western blotting. Bindings among KLF7, HDAC4, microRNA (miR)-148b-3p, and NCOR1 were testified via the dual-luciferase assay and chromatin immunoprecipitation assay. Results: KLF7 was poorly expressed in AS mice and ox-LDL-induced RAW264.7 cells. KLF7 overexpression attenuated AS lesions and rescued metabolic abnormities in AS mice, and reduced glucose intake and GMR in ox-LDL-induced RAW264.7 cells. Mechanically, KLF7 bound to the HDAC4 promoter to activate HDAC4. HDAC4 reduced H3 and H4 acetylation levels in the miR-148b promoter to inhibit miR-148b-3p and promote NCOR1 transcription. HDAC4 downregulation abolished the protective role of KLF7 overexpression in AS mice and ox-LDL-induced RAW264.7 cells via the miR-148b-3p/NCOR1 axis. Conclusion: KLF7 bound to the HDAC4 promoter to activate HDAC4, inhibit miR-148b-3p via reducing acetylation level, and promote NCOR1 transcription, thereby limiting GMR in macrophages and alleviating AS lesions.

Dual Biosignal-Functional Injectable Microspheres for Remodeling Osteogenic Microenvironment.

Bone defect regeneration depends on the population and lifespan of M2 macrophages, which are regulated by dual signals generated by the "physical" spatial configuration of biological tissues and "molecular" chemokines. Herein, inspired by the reprogramming of macrophages, immunoengineered porous microspheres are constructed to accelerate bone repair through the regulation of both "physical" and "molecular" signals. The porous structure of injectable poly (l-lactic acid) (PLLA) microspheres prepared by the microfluidic technique provides a "physical signal" for osteogenic differentiation. Additionally, interleukin (IL)-4-loaded liposomes (Ls) are modified on PLLA microspheres through amide bonds to produce IL-4/Ls/PLLA microspheres, providing a "molecular signal" in stimulating the differentiation of macrophages to M2 type. It is confirmed that IL-4/Ls/PLLA microspheres could induce M2-macrophages polarization and potentiate osteoblast proliferation and differentiation while coculturing with macrophages and osteoblasts in vitro. Besides, IL-4/Ls/PLLA microspheres are proved to promote bone defect regeneration by inducing the conversion of M1 macrophages to M2 through dual biosignal-functional regulation in both the calvaria defect and maxillary sinus defect models. Overall, the immuno-reprogrammed IL-4/Ls/PLLA microspheres achieve the precise immuno-reprogramming of macrophages by dual biosignal-functional regulation. This immune reengineering strategy paves a way for clinical bone defect treatment.

Metabolic reprogramming of tumor-associated macrophages by collagen turnover promotes fibrosis in pancreatic cancer.

A hallmark of pancreatic tumors is their highly desmoplastic stroma composed of fibroblasts, immune cells, and a dense network of collagen fibers. Tumor-associated macrophages are one of the most abundant immune cell populations in the pancreatic tumor stroma. Their protumorigenic function has been attributed predominantly to their capacity to promote immune evasion and metastasis. Tumor-assoc iated macrophages are also well known for their role in the remodeling of the stroma via collagen production and degradation, with the latter being mediated by mannose receptor (MRC1)-dependent endocytosis of collagen. Here we show that MRC1-mediated collagen internalization and subsequent lysosomal degradation by macrophages harboring a tumor-associated phenotype are accompanied by the accumulation of collagen-derived intracellular free amino acids and increased arginine biosynthesis. The resulting increase in intracellular arginine levels leads to the up-regulation of inducible nitric oxide synthase and the production of reactive nitrogen species. Furthermore, reactive nitrogen species derived from internalized and degraded collagen promotes a profibrotic phenotype in pancreatic stellate cells resulting in enhanced intratumoral collagen deposition. Overall, our findings identify a role for extracellular matrix remodeling in the functional modulation of tumor-associated macrophages via metabolic rewiring.

The Secretome of Preconditioned Mesenchymal Stem Cells Drives Polarization and Reprogramming of M2a Macrophages toward an IL-10-Producing Phenotype.

The preconditioning of mesenchymal stem cells (MSCs) has been recognized as an attractive tool to improve their regenerative and immunomodulatory capacities based on their paracrine effects. In this study, we examined the potential of an MSC-conditioned medium (MSC-CM) to alter the phenotype of murine macrophages and to drive reprogramming toward an anti-inflammatory, M2-like state in vitro. We further explored the impact of MSC cytokine preconditioning on the immunosuppressive properties of the MSC secretome. The MSC-CM suppressed the expression of proinflammatory genes in murine M1 macrophages, but only the CM from preconditioned MSCs (preMSC-CM) downregulated their expression during M1 polarization. Remarkably, only the preMSC-CM significantly increased the expression of M2a-, M2b- and M2c-specific genes and proteins during M2a polarization. Further, macrophages were found to secrete high levels of anti-inflammatory IL-10. Similarly, M2a macrophages cultured in the presence of the preMSC-CM displayed an enhanced expression of M2b/M2c-specific markers, suggesting that the secretome of preMSC promotes the repolarization of M2a-like macrophages to M2b/M2c subtypes. The preMSC-CM was found to be enriched in molecules involved in M2 polarization. Additionally, a unique downregulation of extracellular matrix components was observed. Altogether, the preMSC-CM may provide an attractive strategy to dampen inflammation by suppressing the expression of proinflammatory mediators and promoting the polarization and phenotype switch of M2a cells to IL-10-secreting M2b/M2c-like macrophages.

Supramolecular nanotherapeutics enable metabolic reprogramming of tumor-associated macrophages to inhibit tumor growth.

Tumor-associated macrophages (TAMs) exist in multiple phenotypes across the spectrum, defined by an M1 antitumorigenic phenotype and an M2 pro-tumorigenic phenotype on two ends of the spectrum. A largely immunosuppressive tumor-microenvironment aids the polarization of the infiltrating macrophages to a pro-tumorigenic M2 phenotype that promotes tumor progression and metastasis. Recent developments in macrophage immunotherapy have focused on strategies to re-educate TAMs from an M2 to M1 phenotype. Recent findings in the realm of immuno-metabolism have indicated that distinct metabolic signatures accompany macrophages based on their polarization states (M1-Glycolysis and M2-TCA cycle). These metabolites are important drivers of cellular signaling responsible for acquiring these polarization states, with evidence showing that metabolism is essential to facilitate the energy requirements of immune cells and regulate immune cell response. We hypothesized that TAMs could be reprogrammed metabolically by co-delivery of drugs using a supramolecular nanoparticle system that could effectively rewire macrophage metabolism by simultaneous inhibition of the TCA cycle and upregulation of the glycolytic metabolic pathway. TLR7/8 agonist and Fatty Acid Oxidation (FAO) inhibitor loaded metabolic supramolecular nanoparticles (MSNPs) were synthesized. In vitro assays showed macrophages treated with MSNPs were reprogrammed from an M2 phenotype to an M1 phenotype while significantly upregulating phagocytosis. When injected in 4T1 tumor-bearing mice, MSNPs treatment reduced tumor growth progression more than other treatments. Hence, the delivery of TLR7/8 agonist combined with an FAO inhibitor can enhance antitumor efficacy through metabolic reprogramming of tumor-associated macrophages.

MFG-E8 Reduces Aortic Intimal Proliferation in a Murine Model of Transplant Vasculopathy.

Transplant vasculopathy is characterized by endothelial apoptosis, which modulates the local microenvironment. Milk fat globule epidermal growth factor 8 (MFG-E8), which is released by apoptotic endothelial cells, limits tissue damage and inflammation by promoting anti-inflammatory macrophages. We aimed to study its role in transplant vasculopathy using the murine aortic allotransplantation model. BALB/c mice were transplanted with fully mismatched aortic transplants from MFG-E8 knockout (KO) or wild type (WT) C57BL/6J mice. Thereafter, mice received MFG-E8 (or vehicle) injections for 9 weeks prior to histopathological analysis of allografts for intimal proliferation (hematoxylin and eosin staining) and leukocyte infiltration assessment (immunofluorescence). Phenotypes of blood leukocytes and humoral responses were also evaluated (flow cytometry and ELISA). Mice receiving MFG-E8 KO aortas without MFG-E8 injections had the most severe intimal proliferation (p < 0.001). Administration of MFG-E8 decreased intimal proliferation, especially in mice receiving MFG-E8 KO aortas. Administration of MFG-E8 also increased the proportion of anti-inflammatory macrophages among graft-infiltrating macrophages (p = 0.003) and decreased systemic CD4+ and CD8+ T-cell activation (p < 0.001). An increase in regulatory T cells occurred in both groups of mice receiving WT aortas (p < 0.01). Thus, the analarmin MFG-E8 appears to be an important protein for reducing intimal proliferation in this murine model of transplant vasculopathy. MFG-E8 effects are associated with intra-allograft macrophage reprogramming and systemic T-cell activation dampening.

Integrated metabolomic and transcriptomic analysis identifies benzo[a]pyrene-induced characteristic metabolic reprogramming during accumulation of lipids and reactive oxygen species in macrophages

Polycyclic aromatic hydrocarbon exposure is a major risk factor for cardiovascular diseases. Macrophage lipid accumulation is a characteristic molecular event in the pathophysiology of cardiovascular diseases. Metabolic reprogramming is an intervention target for diseases and toxic effects of environmental pollutants. However, comprehensive metabolic reprogramming related to BaP-induced macrophage lipid accumulation is currently unexplored. Therefore, metabolomics and transcriptomics were conducted to unveil relevant metabolic reprogramming in BaP-exposed macrophages, and to discover potential intervention targets. Metabolomics revealed that most amino acids, nucleotides, monosaccharides, and organic acids were significantly decreased, while most fatty acids and steroids accumulated in BaP-exposed macrophages. Transcriptomics showed that fatty acid synthesis and oxidation, and steroid synthesis and export were decreased, while import of fatty acids and steroids was increased, indicating potential roles of lipid transport in macrophage lipid accumulation following BaP exposure. Meanwhile, alanine, aspartate and glutamate metabolism, branched-chain amino acid degradation, nucleotide synthesis, monosaccharide import, pentose phosphate pathway, citrate synthesis, and glycolysis were decreased, while nucleotide degradation was increased, thus inducing decreases in most amino acids, nucleotides, monosaccharides, and organic acids in BaP-exposed macrophages. Additionally, increases in oxidative stress and the activation of antioxidant systems were observed in BaP-exposed macrophages, which was evinced by increases in reactive oxygen species, and the activation of Fenton reaction, Vdac2/3, Sod2, and Nrf2. Moreover, BaP-induced accumulation of reactive oxygen species and lipids in macrophages could be abolished by epigallocatechin-3-gallate. Quantitative PCR showed that BaP exposure activated aryl hydrocarbon receptor signaling and promoted the proinflammatory phenotype in macrophages, and these effects were inhibited or even abolished by the separate treatment with epigallocatechin-3-gallate or CH-223191, suggesting the regulatory role of aryl hydrocarbon receptor signaling in BaP-induced toxic effects. This study provides novel insights into the toxic effects of polycyclic aromatic hydrocarbons on macrophage metabolism and potential intervention targets.

Chlorin e6-Induced Photodynamic Effect Polarizes the Macrophage Into an M1 Phenotype Through Oxidative DNA Damage and Activation of STING.

The tumor-associated macrophage (TAM) serves as an immunosuppressive agent in the malignant tumor microenvironment, facilitating the development and metastasis of lung cancer. The photodynamic effect destabilizes cellular homeostasis owing to the generation of reactive oxygen species (ROS), resulting in the enhanced pro-inflammatory function of immunocytes. In our previous study, the Ce6-mediated photodynamic effect was found to have kept the viability of macrophages and to remodel them into the M1 phenotype. However, the mechanism remains unrevealed. The present study now explores the mechanism of photodynamic therapy (PDT)-mediated reprogramming of macrophages. As expected, Ce6-mediated PDT was capable of generating reactive oxygen species, which was continuously degraded, causing “low intensity” damage to DNA and thereby triggering subsequent DNA damage response in macrophages. The autophagy was thus observed in Ce6-treated macrophages and was shown to protect cells from being photodynamically apoptotic. More importantly, Ce6 PDT could activate the stimulator of interferon genes (STING) molecule, a sensor of DNA damage, which could activate the downstream nuclear factor kappa-B (NF-κB) upon activation, mediating the polarization of macrophages towards the M1 phenotype thereupon. In addition, inhibition of ROS induced by PDT attenuated the DNA damage, STING activation, and M1-phenotype reprogramming. Furthermore, the silence of the STING weakened Ce6 treatment-mediated M1 remodeling of macrophages as well. Altogether, these findings indicate the Ce6-induced photodynamic effect polarizes macrophages into an M1 phenotype through oxidative DNA damage and subsequent activation of the STING. This work reveals the crucial mechanism by which photodynamic therapy regulates the macrophage phenotype and also provides a novel intervenable signaling target for remodeling macrophages into the M1 phenotype.

An imbalance of the IL-33/ST2-AXL-efferocytosis axis induces pregnancy loss through metabolic reprogramming of decidual macrophages.

During embryo implantation, apoptosis is inevitable. These apoptotic cells (ACs) are removed by efferocytosis, in which macrophages are filled with a metabolite load nearly equal to the phagocyte itself. A timely question pertains to the relationship between efferocytosis-related metabolism and the immune behavior of decidual macrophages (dMΦs) and its effect on pregnancy outcome. Here, we report positive feedback of IL-33/ST2-AXL-efferocytosis leading to pregnancy failure through metabolic reprogramming of dMΦs. We compared the serum levels of IL-33 and sST2, along with IL-33 and ST2, efferocytosis and metabolism of dMΦs, from patients with normal pregnancies and unexplained recurrent pregnancy loss (RPL). We revealed disruption of the IL-33/ST2 axis, increased apoptotic cells and elevated efferocytosis of dMΦs from patients with RPL. The dMΦs that engulfed many apoptotic cells secreted more sST2 and less TGF-β, which polarized dMΦs toward the M1 phenotype. Moreover, the elevated sST2 biased the efferocytosis-related metabolism of RPL dMΦs toward oxidative phosphorylation and exacerbated the disruption of the IL-33/ST2 signaling pathway. Metabolic disorders also lead to dysfunction of efferocytosis, resulting in more uncleared apoptotic cells and secondary necrosis. We also screened the efferocytotic molecule AXL regulated by IL-33/ST2. This positive feedback axis of IL-33/ST2-AXL-efferocytosis led to pregnancy failure. IL-33 knockout mice demonstrated poor pregnancy outcomes, and exogenous supplementation with mouse IL-33 reduced the embryo losses. These findings highlight a new etiological mechanism whereby dMΦs leverage immunometabolism for homeostasis of the microenvironment at the maternal-fetal interface.

PERK is a critical metabolic hub for immunosuppressive function in macrophages.

Chronic inflammation triggers compensatory immunosuppression to stop inflammation and minimize tissue damage. Studies have demonstrated that endoplasmic reticulum (ER) stress augments the suppressive phenotypes of immune cells; however, the molecular mechanisms underpinning this process and how it links to the metabolic reprogramming of immunosuppressive macrophages remain elusive. In the present study, we report that the helper T cell 2 cytokine interleukin-4 and the tumor microenvironment increase the activity of a protein kinase RNA-like ER kinase (PERK)-signaling cascade in macrophages and promote immunosuppressive M2 activation and proliferation. Loss of PERK signaling impeded mitochondrial respiration and lipid oxidation critical for M2 macrophages. PERK activation mediated the upregulation of phosphoserine aminotransferase 1 (PSAT1) and serine biosynthesis via the downstream transcription factor ATF-4. Increased serine biosynthesis resulted in enhanced mitochondrial function and α-ketoglutarate production required for JMJD3-dependent epigenetic modification. Inhibition of PERK suppressed macrophage immunosuppressive activity and could enhance the efficacy of immune checkpoint programmed cell death protein 1 inhibition in melanoma. Our findings delineate a previously undescribed connection between PERK signaling and PSAT1-mediated serine metabolism critical for promoting immunosuppressive function in M2 macrophages.

Dihydronicotinamide Riboside Is a Potent NAD+ Precursor Promoting a Pro-Inflammatory Phenotype in Macrophages

Nicotinamide adenine dinucleotide (NAD) metabolism plays an important role in the regulation of immune function. However, a complete picture of how NAD, its metabolites, precursors, and metabolizing enzymes work together in regulating immune function and inflammatory diseases is still not fully understood. Surprisingly, few studies have compared the effect of different forms of vitamin B3 on cellular functions. Therefore, we investigated the role of NAD boosting in the regulation of macrophage activation and function using different NAD precursors supplementation. We compared nicotinamide mononucleotide (NMN), nicotinamide riboside (NR), and nicotinamide (NAM) supplementation, with the recently described potent NAD precursor NRH. Our results show that only NRH supplementation strongly increased NAD+ levels in both bone marrow-derived and THP-1 macrophages. Importantly, NRH supplementation activated a pro-inflammatory phenotype in resting macrophages, inducing gene expression of several cytokines, chemokines, and enzymes. NRH also potentiated the effect of lipopolysaccharide (LPS) on macrophage activation and cytokine gene expression, suggesting that potent NAD+ precursors can promote inflammation in macrophages. The effect of NRH in NAD+ boosting and gene expression was blocked by inhibitors of adenosine kinase, equilibrative nucleoside transporters (ENT), and IκB kinase (IKK). Interestingly, the IKK inhibitor, BMS-345541, blocked the mRNA expression of several enzymes and transporters involved in the NAD boosting effect of NRH, indicating that IKK is also a regulator of NAD metabolism. In conclusion, NAD precursors such as NRH may be important tools to understand the role of NAD and NADH metabolism in the inflammatory process of other immune cells, and to reprogram immune cells to a pro-inflammatory phenotype, such as the M2 to M1 switch in macrophage reprogramming, in the cancer microenvironment.

Exosome-mediated genetic reprogramming of tumor-associated macrophages by exoASO-STAT6 leads to potent monotherapy antitumor activity.

Effectiveness of checkpoint immunotherapy in cancer can be undermined by immunosuppressive tumor-associated macrophages (TAMs) with an M2 phenotype. Reprogramming TAMs toward a proinflammatory M1 phenotype is a novel approach to induce antitumor immunity. The M2 phenotype is controlled by key transcription factors such as signal transducer and activator of transcription 6 (STAT6), which have been “undruggable” selectively in TAMs. We describe an engineered exosome therapeutic candidate delivering an antisense oligonucleotide (ASO) targeting STAT6 (exoASO-STAT6), which selectively silences STAT6 expression in TAMs. In syngeneic models of colorectal cancer and hepatocellular carcinoma, exoASO-STAT6 monotherapy results in >90% tumor growth inhibition and 50 to 80% complete remissions. Administration of exoASO-STAT6 leads to induction of nitric oxide synthase 2 (NOS2), an M1 macrophage marker, resulting in remodeling of the tumor microenvironment and generation of a CD8 T cell–mediated adaptive immune response. Collectively, exoASO-STAT6 represents the first platform targeting transcription factors in TAMs in a highly selective manner.

Abstract P5-16-02: Updated efficacy, safety and translational data from MARIO-3, a phase II open-label study evaluating a novel triplet combination of eganelisib (IPI-549), atezolizumab (atezo), and nab-paclitaxel (nab-pac) as first-line (1L) therapy for locally advanced or metastatic triple-negative breast cancer (TNBC)

Abstract Background: Eganelisib is a first-in-class, oral, immuno-oncology macrophage reprogramming agent. The IMpassion130 randomized trial in 1L advanced TNBC has demonstrated improved efficacy with the addition of atezo to nab-pac in patients with PD-L1+ tumors. Adding eganelisib to the approved doublet of atezo and nab-pac is expected to improve patient benefit regardless of PD-L1 status. Methods: Eligible patients (pts) had measurable unresectable locally advanced or metastatic TNBC and no prior systemic therapy for advanced disease. A safety run-in assessed the safety of the triplet of oral eganelisib 30 mg daily in combination with IV nab-pac 100 mg/m2 given on days 1, 8, &amp; 15, and IV atezo 840 mg given on days 1 &amp; 15 of a 28 day cycle. After establishing tolerability and confirmation of the 30 mg dose in the safety run-in (n=6), the expansion phase of the phase II study was initiated to enroll approximately 60 pts (30 PD-L1+ and 30 PD-L1-). Cycles are repeated until loss of clinical benefit, unacceptable toxicity or consent withdrawal. The primary efficacy endpoint is confirmed complete response (CR) rate per RECIST v1.1. Secondary endpoints include the overall response rate (ORR), progression free survival and safety assessment. Results: As of 6/26/2021, data from the safety run-in and the expansion cohorts are reported for 43 pts evaluable for safety and 38 evaluable for efficacy defined as having at least one post-baseline tumor assessment. 2 CRs (2/38), 19 PRs (19/38), and 11 SDs (11/38) were observed with an ORR of 55.3% (21/38) and a disease control rate (DCR) of 84.2% (32/38). Responses were seen irrespective of PD-L1 status; in PD-L1+ pts, ORR is 66.7% (8/12) and DCR is 91.7% (11/12), and in PD-L1- pts, ORR is 47.8% (11/23) and DCR is 78.3% (18/23). The most common all-grade adverse events were nausea (51.2%), fatigue (48.8%), alopecia (32.6%), diarrhea (32.6%), rash maculo-papular (30.2%), ALT increased (27.9%), and AST increased (25.6%). The most common grade ≥3 adverse events were ALT increased (18.6%), AST increased (14.0%), neutrophil count decreased (9.3%) and rash maculo-papular (9.3%). The safety profile is acceptable and in line with expectations of component drugs with no additive or new safety signals. Translational data from paired biopsies show a shift in tumor associated macrophages towards an immune activated state as well as an increase in cytotoxic T cells. In addition, a subset of tumors with baseline PD-L1 negative status show conversion to PD-L1 positive status post treatment. Consistent with immune activation, increased interferon gamma signature, increased proinflammatory cytokines and T cell invigoration were observed in peripheral blood cells. Conclusions: The novel triplet regimen of eganelisib, atezo, and nab-pac shows promising anti-tumor activity (ORR 55.3% and DCR 84.2%) irrespective of PD-L1 status and has manageable toxicity. . Translational data from paired biopsies and peripheral blood show evidence of immune activation regardless of PD-L1 status as well as conversion from PD-L1 negative to PD-L1 positive subtype within a subset of patients treated with the triplet regimen. The expansion phase of the phase II study is currently enrolling. At the time of presentation, we will have updated efficacy, safety and translational data. Citation Format: Soliman Hatem, Jeffrey Hargis, Anthony Elias, Arielle Lee, Rachel Swart, Shaker Dahkil, Alexandra Drakaki, Vu Phan, Frederic Kass, Melody Cobleigh, Sanil Babu, Katherine Tkaczuk, Brenda O'Connell, Jennifer Roberts, Nora Zizlsperger, Erika Hamilton. Updated efficacy, safety and translational data from MARIO-3, a phase II open-label study evaluating a novel triplet combination of eganelisib (IPI-549), atezolizumab (atezo), and nab-paclitaxel (nab-pac) as first-line (1L) therapy for locally advanced or metastatic triple-negative breast cancer (TNBC) [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-16-02.

Monocyte and Macrophage Lipid Accumulation Results in Down-Regulated Type-I Interferon Responses.

Macrophages are critical components of atherosclerotic lesions and their pro- and anti-inflammatory responses influence atherogenesis. Type-I interferons (IFNs) are cytokines that play an essential role in antiviral responses and inflammatory activation and have been shown to promote atherosclerosis. Although the impact of type-I IFNs on macrophage foam cell formation is well-documented, the effect of lipid accumulation in monocytes and macrophages on type-I IFN responses remains unknown. Here we examined IFN stimulated (ISG) and non-ISG inflammatory gene expression in mouse and human macrophages that were loaded with acetylated LDL (acLDL), as a model for foam cell formation. We found that acLDL loading in mouse and human macrophages specifically suppressed expression of ISGs and IFN-β secretion, but not other pro-inflammatory genes. The down regulation of ISGs could be rescued by exogenous IFN-β supplementation. Activation of the cholesterol-sensing nuclear liver X receptor (LXR) recapitulated the cholesterol-initiated type-I IFN suppression. Additional analyses of murine in vitro and in vivo generated foam cells confirmed the suppressed IFN signaling pathways and suggest that this phenotype is mediated via down regulation of interferon regulatory factor binding at gene promoters. Finally, RNA-seq analysis of monocytes of familial hypercholesterolemia (FH) patients also showed type-I IFN suppression which was restored by lipid-lowering therapy and not present in monocytes of healthy donors. Taken together, we define type-I IFN suppression as an athero-protective characteristic of foamy macrophages. These data provide new insights into the mechanisms that control inflammatory responses in hyperlipidaemic settings and can support future therapeutic approaches focusing on reprogramming of macrophages to reduce atherosclerotic plaque progression and improve stability.

MiR-182 targeting reprograms tumor-associated macrophages and limits breast cancer progression.

The protumor roles of alternatively activated (M2) tumor-associated macrophages (TAMs) have been well established, and macrophage reprogramming is an important therapeutic goal. However, the mechanisms of TAM polarization remain incompletely understood, and effective strategies for macrophage targeting are lacking. Here, we show that miR-182 in macrophages mediates tumor-induced M2 polarization and can be targeted for therapeutic macrophage reprogramming. Constitutive miR-182 knockout in host mice and conditional knockout in macrophages impair M2-like TAMs and breast tumor development. Targeted depletion of macrophages in mice blocks the effect of miR-182 deficiency in tumor progression while reconstitution of miR-182-expressing macrophages promotes tumor growth. Mechanistically, cancer cells induce miR-182 expression in macrophages by TGFβ signaling, and miR-182 directly suppresses TLR4, leading to NFκb inactivation and M2 polarization of TAMs. Importantly, therapeutic delivery of antagomiR-182 with cationized mannan-modified extracellular vesicles effectively targets macrophages, leading to miR-182 inhibition, macrophage reprogramming, and tumor suppression in multiple breast cancer models of mice. Overall, our findings reveal a crucial TGFβ/miR-182/TLR4 axis for TAM polarization and provide rationale for RNA-based therapeutics of TAM targeting in cancer.

Polypharmacologic Reprogramming of Tumor-Associated Macrophages toward an Inflammatory Phenotype.

Tumor-associated macrophages (TAM) are an important component of the tumor microenvironment (TME) that can promote tumor progression, metastasis, and resistance to therapies. Although TAMs represent a promising target for therapeutic intervention, the complexity of the TME has made the study of TAMs challenging. Here, we established a physiologically relevant in vitro TAM polarization system that recapitulates TAM protumoral activities. This system was used to characterize dynamic changes in gene expression and protein phosphorylation during TAM polarization and to screen phenotypic kinase inhibitors that impact TAM programming. BMS-794833, a multitargeted compound, was identified as a potent inhibitor of TAM polarization. BMS-794833 decreased protumoral properties of TAMs in vitro and suppressed tumor growth in mouse triple-negative breast cancer models. The effect of BMS-794833 was independent of its primary targets (MET and VEGFR2) but was dependent on its effect on multiple signaling pathways, including focal adhesion kinases, SRC family kinases, STAT3, and p38 MAPKs. Collectively, these findings underline the efficacy of polypharmacologic strategies in reprogramming complex signaling cascades activated during TAM polarization. SIGNIFICANCE: A physiologically relevant in vitro system of TAM polarization uncovers signaling pathways that regulate polarization and identifies strategies to target macrophage reprogramming to suppress cancer growth.

MARCKS/AXL Fibrogenic Complex Activated in Tobacco Smoke Promotes Macrophage Reprogramming and Pulmonary Fibrosis

MARCKS/AXL Fibrogenic Complex Activated in Tobacco Smoke Promotes Macrophage Reprogramming and Pulmonary Fibrosis

Abu Khaled Muhammed Akramul Haq

<h3>Rationale</h3> A better understanding of the mechanism of action of mesenchymal stromal cells (MSCs) and their extracellular vesicles (EVs) is needed to support their use as novel therapies for acute...

Self-assembled tetrahedral framework nucleic acid mediates tumor-associated macrophage reprogramming and restores antitumor immunity

There is increasing interest in depleting or repolarizing tumor-associated macrophages (TAMs) to generate a proinflammatory effect. However, TAMs usually display an immunosuppressive M2-like phenotype in the tumor microenvironment. Apparently, developing a macrophage-targeting delivery system with immunomodulatory agents is urgent. In this study, an efficient siRNA and CpG ODNs delivery system (CpG-siRNA-tFNA) was prepared with nucleic acid stepwise self-assembled. The tFNA composed of CpG ODNs and siRNA showed a higher stability and an enhanced cellular uptake efficiency. Moreover, the CpG-siRNA-tFNA effectively reprogrammed TAMs toward M1 phenotype polarization with increased proinflammatory cytokine secretion and NF-κB signal pathway activation, which triggers dramatic antitumor immune responses. Additionally, the CpG-siRNA-tFNA exhibited superior antitumor efficacy in a breast cancer xenograft mouse model without obvious systemic side effects. Taken together, CpG-siRNA-tFNA displayed greatly antitumor effect by facilitating TAM polarization toward M1 phenotypes in favor of immunotherapy. Hence, we have developed an efficient therapeutic strategy with immunomodulatory agents for clinical applications.