Search Results (13)

Background/Objectives: CD47 is a cell surface glycoprotein moderately expressed in healthy cells and upregulated in cancer and viral infected cells. CD47’s interaction with signal regulatory protein alpha (SIRPα) inhibits phagocytic cells and its interaction with thrombospondin-1 inhibits T cell response. Experimental evidence has revealed that the blockade of CD47 resulted in the increased activation and function of both innate and adaptive immune cells, therefore exerting antitumoral and antiviral effects. Recent studies have shown that the combination of vaccines and immune checkpoint inhibitors could be a promising approach to increasing vaccine immunogenicity. Here, we investigated the vaccinal effect of anti-CD47 antibodies and discussed the possibilities of combining anti-CD47 treatments with vaccines. Methods: Using vesicular stomatitis virus (VSV), a widely used replication-competent vaccine vector, we evaluated the impact of the immunotherapeutic blockade of CD47 on cellular, humoral, and protective immunity. We infected C57BL/6 mice with VSV, treated them with anti-CD47 antibodies or an isotype, and evaluated the total immunoglobulin (Ig), IgG neutralizing antibodies, B cell activation, CD8+ T cell effector function, and survival of the mice. Results: We found that the treatments of anti-CD47 antibodies led to significantly increased Ig and IgG neutralizing antibody levels compared to the isotype treatment. Flow cytometric analysis of B cells revealed no difference in the number of circulating B cells; however, we observed an increased surface expression of CD80 and CD86 in B cells among anti-CD47-treated mice. Further analysis of the impact of CD47 blockade on T immunity revealed a significantly higher percentage of IFN-γ⁺ CD4 and IFN-γ⁺ CD8 T cells in anti-CD47-treated mice. Upon infecting mice with a lethal VSV dose, we observed a significantly higher survival rate among the anti-CD47-treated mice compared to control mice. Conclusions: Our results indicate that anti-CD47 treatment induces a stronger cellular and humoral immune response, leading to better protection. As such, immunotherapy by CD47 blockade in combination with vaccines could be a promising approach to improve vaccine efficacy. Full article

Background: Macrophages play an important role in eliminating diseased and damaged cells through programmed cell death. Signal regulatory protein alpha (SIRPα) is a crucial immune checkpoint primarily expressed on myeloid cells and macrophages. It initiates a ‘do not eat me’ signal when engaged with CD47, which is typically expressed at elevated levels on multiple solid tumors. The phospholipase A2 Group 7 (PLA2G7), which is mainly secreted by macrophages, interacts with oxidized low-density lipoprotein (oxLDL) and associates with several vascular diseases and cancers. Methods: To identify potent fully human monoclonal antibodies (mAbs) against human SIRPα and PLA2G7, we conducted bio-panning of phage antibody libraries. Results: We isolated one human Fab (1B3) and VH (1A3) for SIRPα, as well as one human Fab (1H8) and one VH (1A9) for PLA2G7; the 1B3 Fab and 1A3 VH are competitively bound to SIRPα, interfering with CD47 binding. The 1B3 IgG and 1A3 VH-Fc augmented macrophage-mediated phagocytic activity when combined with the anti-EGFR antibody, cetuximab. The anti-PLA2G7 antibodies exhibited high specificity for the PLA2G7 antigen and effectively blocked the PLA2G7 enzymatic activity with half-maximal inhibitory concentrations (IC₅₀) in the single-digit nanomolar range. Additionally, 1H8 IgG and its derivative bispecific antibody exhibited the ability to block PLA2G7-mediated tumor cell migration. Conclusions: Our anti-SIRPα mAbs are expected to serve as potent and fully human immune checkpoint inhibitors of SIRPα, enhancing the antitumor responses of SIRPα-positive immune cells. Moreover, our anti-PLA2G7 mAbs represent promising fully human PLA2G7 enzymatic blockade antibodies with the potential to enhance both anti-tumor and anti-aging responses. Anti-SIRPα and PLA2G7 mAbs can modulate macrophage phagocytic activity and inflammatory responses against tumors. Full article

The liver, as the largest metabolic and detoxification organ in mammals, metabolizes approximately 80–90% of drugs. However, drug-induced liver injury (DILI) is common and driven by factors such as individual variability, differences in liver metabolism, and improper drug use. Mesenchymal stem cells (MSCs), with their self-renewal and multipotent differentiation capabilities, offer therapeutic potential, but face challenges such as limited proliferation and increased apoptosis during in vitro expansion. Although MSCs exhibit low immunogenicity, they are often cleared by the host immune system, which limits their survival and engraftment. Glutathione peroxidase 3 (GPX3) is a key antioxidant enzyme that reduces reactive oxygen species (ROS), protecting cells from oxidative damage. CD47, also known as integrin-associated protein (IAP), helps cells evade immune clearance by binding to signal regulatory protein alpha (SIRPα) on the immune cells. Here, we used an acetaminophen (APAP)-induced DILI mouse model to evaluate the therapeutic efficacy of intravenously infused MSCs overexpressing GPX3 and CD47. Compared to unmodified MSCs, modified MSCs showed improved survival, reduced liver inflammation, and alleviated oxidative damage, offering enhanced protection against APAP-induced DILI. Full article

The intricate complex system of the differentiation 47 (CD47) and the signal-regulatory protein alpha (SIRPα) cluster is a crucial target for cancer immunotherapy. Although the conformational state of the CD47-SIRPα complex has been revealed through crystallographic studies, further characterization is needed to fully understand the binding mechanism and to identify the hot spot residues involved. In this study, molecular dynamics (MD) simulations were carried out for the complexes of CD47 with two SIRPα variants (SIRPαv1, SIRPαv2) and the commercially available anti-CD47 monoclonal antibody (B6H12.2). The calculated binding free energy of CD47-B6H12.2 is lower than that of CD47-SIRPαv1 and CD47-SIRPαv2 in all the three simulations, indicating that CD47-B6H12.2 has a higher binding affinity than the other two complexes. Moreover, the dynamical cross-correlation matrix reveals that the CD47 protein shows more correlated motions when it binds to B6H12.2. Significant effects were observed in the energy and structural analyses of the residues (Glu35, Tyr37, Leu101, Thr102, Arg103) in the C strand and FG region of CD47 when it binds to the SIRPα variants. The critical residues (Leu30, Val33, Gln52, Lys53, Thr67, Arg69, Arg95, and Lys96) were identified in SIRPαv1 and SIRPαv2, which surround the distinctive groove regions formed by the B2C, C’D, DE, and FG loops. Moreover, the crucial groove structures of the SIRPα variants shape into obvious druggable sites. The C’D loops on the binding interfaces undergo notable dynamical changes throughout the simulation. For B6H12.2, the residues Tyr32_LC, His92_LC, Arg96_LC, Tyr32_HC, Thr52_HC, Ser53_HC, Ala101_HC, and Gly102_HC in its initial half of the light and heavy chains exhibit obvious energetic and structural impacts upon binding with CD47. The elucidation of the binding mechanism of SIRPαv1, SIRPαv2, and B6H12.2 with CD47 could provide novel perspectives for the development of inhibitors targeting CD47-SIRPα. Full article

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

M1 macrophages are an important cell type related to tumor immunology and are known to phagocytose cancer cells. In previous studies, the organogermanium compound poly-trans-[(2-carboxyethyl)germasesquioxane] (Ge-132) and its hydrolysate, 3-(trihydroxygermyl) propanoic acid (THGP), have been reported to exert antitumor effects by activating NK cells and macrophages through the induction of IFN-γ activity in vivo. However, the detailed molecular mechanism has not been clarified. In this study, we found that macrophages differentiate into the M1 phenotype via NF-κB activation under long-term culture in the presence of THGP in vitro and in vivo. Furthermore, long-term culture with THGP increases the ability of RAW 264.7 cells to suppress B16 4A5 melanoma cell proliferation. These mechanisms indicate that THGP promotes the M1 polarization of macrophages and suppresses the expression of signal-regulatory protein alpha (SIRP-α) in macrophages and CD47 in cancers. Based on these results, THGP may be considered a new regulatory reagent that suppresses tumor immunity. Full article

Background and Aims: The macrophage “don’t eat me” pathway CD47/SIRPα is a target for promising new immunotherapy. We hypothesized that a soluble variant of SIRPα is present in the blood and may function as a biomarker. Methods: Monocyte derived macrophages (MDMs) from human buffy-coats were stimulated into macrophage subtypes by LPS and IFN-γ (M1), IL-4 and IL-13 (M2a), IL-10 (M2c) and investigated using flow cytometry. Soluble SIRPα (sSIRPα) was measured in cell cultures and serum by Western blotting and an optimized ELISA. Serum samples were obtained from 120 healthy individuals and from 8 individuals challenged by an LPS injection. Results: All macrophage phenotypes expressed SIRPα by flowcytometry, and sSIRPα was present in all culture supernatants including unstimulated cells. M1 macrophages expressed the lowest level of SIRPαand released the highest level of sSIRPα (p < 0.05). In vivo, the serum level of sSIRPα increased significantly (p < 0.0001) after an LPS challenge in humans. The median concentration in healthy individuals was 28.7 µg/L (19.8–41.1, 95% reference interval), and 20.5 µg/L in an IFCC certified serum reference material. The protein was stable in serum for prolonged storage and repeated freeze/thawing. Conclusions: We demonstrate that sSIRPα is produced constitutively and the concentration increases upon macrophage activation both in vitro and in vivo. It is present in human serum where it may function as a biomarker for the activity of tumor-associated macrophages (TAMs), and for monitoring the effect of immunotherapy. Full article

Background: Cancer cells escape macrophage phagocytosis by expressing the CD47 integrin-associated protein that binds to the SIRPα ligand (signal regulatory protein alpha) expressed by macrophages. Immunotherapy targeting this pathway is under clinical development. Methods: We investigated the expression of CD47/SIRPα molecules in a series of 98 NSCLCs, in parallel with the infiltration of tumor stroma by CD68+ macrophages, tumor-infiltrating lymphocytes (TILs), and PD-L1/PD-1 molecules. Results: Extensive membranous CD47 expression by cancer cells characterized 29/98 cases. SIRPα and CD68 were expressed, to a varying extent, by tumor-associated macrophages (Μφ, TAMs). A high CD68Mφ-score in inner tumor areas was linked with improved overall survival (p = 0.005); and this was independent of the stage (p = 0.02, hazard ratio 0.4). In contrast, high SIRPα expression by CD68+ TAMs (SIRPα/CD68-ratio) was linked with CD47 expression by cancer cells, low TIL-score, and poor prognosis (p = 0.02). A direct association of CD47 expression by cancer cells and the % FOXP3+ TILs (p = 0.01, r = 0.25) was also noted. Conclusions: TAMs play an important role in the prognosis of operable NSCLC. As SIRPα+ macrophages adversely affect prognosis, it is suggested that the CD47/SIRPα axis is a sound target for adjuvant immunotherapy policies, aiming to improve the cure rates in operable NSCLC. Full article

CD47 is a surface membrane protein expressed by all normal tissues. It is the so-called “don’t eat me signal” because it protects the cells against phagocytosis. The CD47 interacts with the signal regulatory protein alpha (SIRPα) on the surface of macrophages, leading to downstream inhibitory signaling that dampens phagocytic capacity. Since macrophages exert immune surveillance against cancers, cancer cells overexpress CD47 to defend themselves against phagocytosis. Acute myeloid leukemia (AML) is a cancer of hematopoietic stem/progenitor cells (HSPC), and similar to other types of cancers, leukemic blasts show enhanced levels of CD47. In patients with AML, CD47 has been associated with a higher disease burden and poor overall survival. Blockage of CD47-SIRPα signaling leads to improved phagocytosis of AML cells and better overall survival in xenograft models. However, the introduction of a pro-phagocytic signal is needed to induce greater phagocytic capacity. These pro-phagocytic signals can be either Fc receptor stimulants (such as monoclonal antibodies) or natural pro-phagocytic molecules (such as calreticulin). Based on these pre-clinical findings, various clinical trials investigating the blockade of CD47-SIRPα interaction have been designed as monotherapy and in combination with other anti-leukemic agents. In this review, we will discuss CD47 biology, highlight its implications for AML pathophysiology, and explore the potential clinical translation of disrupting CD47-SIRPα to treat patients with AML. Full article

Background: To investigate the role of CD47 in inflammatory responses in systemic lupus erythematosus (SLE). Methods: Expression of CD47 and signal regulatory protein alpha (SIRPα) by peripheral blood mononuclear cells (PBMCs) and changes in CD47 expression after exposure to SLE serum, healthy control (HC) serum, recombinant interferon (IFN)-α, or tumor necrosis factor (TNF)-α were examined. Human monocytes and THP1 cells were incubated with lipopolysaccharide (LPS), an anti-CD47 antibody, or both. TNF-α production was examined. Sera from SLE patients and HCs were screened to detect autoantibodies specific for CD47. Results: Twenty-five SLE patients and sixteen HCs were enrolled. CD47 expression by monocytes from SLE patients was higher than those from HCs (mean fluorescence intensity ± SD: 815.9 ± 269.4 vs. 511.5 ± 199.4, respectively; p < 0.001). CD47 expression by monocytes correlated with SLE disease activity (Spearman’s rho = 0.467, p = 0.019). IFN-α but not TNF-α, increased CD47 expression. Exposing monocytes to an anti-CD47 antibody plus LPS increased TNF-α production by 21.0 ± 10.9-fold (compared with 7.3 ± 5.5-fold for LPS alone). Finally, levels of autoantibodies against CD47 were higher in SLE patients than in HCs (21.4 ± 7.1 ng/mL vs. 16.1 ± 3.1 ng/mL, respectively; p = 0.02). Anti-CD47 antibody levels did not correlate with disease activity (Spearman’s rho = −0.11, p = 0.759) or CD47 expression on CD14 monocytes (Spearman’s rho = 0.079, p = 0.838) in patients. Conclusions: CD47 expression by monocytes is upregulated in SLE and correlates with disease activity. CD47 contributes to augmented inflammatory responses in SLE. Targeting CD47 might be a novel treatment for SLE. Full article

Non-thermal plasma (NTP) therapy has been emerging as a promising cancer treatment strategy, and recently, its ability to locally induce immunogenic cancer cell death is being unraveled. We hypothesized that the chemical species produced by NTP reduce immunosuppressive surface proteins and checkpoints that are overexpressed on cancerous cells. Here, 3D in vitro tumor models, an in vivo mouse model, and molecular dynamics simulations are used to investigate the effect of NTP on CD47, a key innate immune checkpoint. CD47 is immediately modulated after NTP treatment and simulations reveal the potential oxidized salt-bridges responsible for conformational changes. Umbrella sampling simulations of CD47 with its receptor, signal-regulatory protein alpha (SIRPα), demonstrate that the induced-conformational changes reduce its binding affinity. Taken together, this work provides new insight into fundamental, chemical NTP-cancer cell interaction mechanisms and a previously overlooked advantage of present NTP cancer therapy: reducing immunosuppressive signals on the surface of cancer cells. Full article

Background: Pancreatic ductal adenocarcinoma (PDAC) is resistant to single-agent immunotherapies. To understand the mechanisms leading to the poor response to this treatment, a better understanding of the PDAC immune landscape is required. The present work aims to study the immune profile in PDAC in relationship to spatial heterogeneity of the tissue microenvironment (TME) in intact tissues. Methods: Serial section and multiplex in situ analysis were performed in 42 PDAC samples to assess gene and protein expression at single-cell resolution in the: (a) tumor center (TC), (b) invasive front (IF), (c) normal parenchyma adjacent to the tumor, and (d) tumor positive and negative draining lymph nodes (LNs). Results: We observed: (a) enrichment of T cell subpopulations with exhausted and senescent phenotype in the TC, IF and tumor positive LNs; (b) a dominant type 2 immune response in the TME, which is more pronounced in the TC; (c) an emerging role of CD47-SIRPα axis; and (d) a similar immune cell topography independently of the neoadjuvant chemotherapy. Conclusion: This study reveals the existence of dysfunctional T lymphocytes with specific spatial distribution, thus opening a new dimension both conceptually and mechanistically in tumor-stroma interaction in PDAC with potential impact on the efficacy of immune-regulatory therapeutic modalities. Full article

Tissue contacting surfaces of medical devices initiate a host inflammatory response, characterized by adsorption of blood proteins and inflammatory cells triggering the release of cytokines, reactive oxygen species (ROS) and reactive nitrogen species (RNS), in an attempt to clear or isolate the foreign object from the body. This normal host response contributes to device-associated pathophysiology and addressing device biocompatibility remains an unmet need. Although widespread attempts have been made to render the device surfaces unreactive, the establishment of a completely bioinert coating has been untenable and demonstrates the need to develop strategies based upon the molecular mechanisms that define the interaction between host cells and synthetic surfaces. In this review, we discuss a family of transmembrane receptors, known as immunoreceptor tyrosine-based inhibitory motif (ITIM)-containing receptors, which show promise as potential targets to address aberrant biocompatibility. These receptors repress the immune response and ensure that the intensity of an immune response is appropriate for the stimuli. Particular emphasis will be placed on the known ITIM-containing receptor, Signal Regulatory Protein Alpha (SIRPα), and its cognate ligand CD47. In addition, this review will discuss the potential of other ITIM-containing proteins as targets for addressing the aberrant biocompatibility of polymeric biomaterials. Full article