Anti-mouse CD40 Monoclonal Antibody (Clone: FGK4.5) | PA007274.r2a

Catalog No.	PA007274.r2a
Product Name	Anti-mouse CD40 Monoclonal Antibody (Clone: FGK4.5) | PA007274.r2a
Supplier Name	Syd Labs, Inc.
Brand Name	Syd Labs
Synonyms	cluster of differentiation 40, Bp50, CDW40, TNFRSF5, p50
Summary	The anti-mouse CD40 monoclonal antibody (clone: FGK4.5) was produced in mammalian cells.
Clone	FGK4.5 / FGK45
Isotype	Rat IgG2a Kappa
Specificity/Sensitivity	The in vivo grade recombinant rat monoclonal antibody (clone: FGK4.5) specifically binds to the mouse CD40 protein.
Applications	ELISA, flow cytometry, neutralization, functional assays such as bioanalytical PK and ADA assays, and those assays for studying biological pathways affected by the mouse CD40 protein.
Form Of Antibody	0.2 uM filtered solution, pH 7.4, no stabilizers or preservatives.
Endotoxin	< 1 EU per 1 mg of the protein by the LAL method.
Purity	>95% by SDS-PAGE under reducing conditions and HPLC.
Shipping	The antibody is shipped with ice pack. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 12 months from date of receipt, -20 to -70°C as supplied. 1 month from date of receipt, 2 to 8°C as supplied.
Note	Recombinant rat IgG2a isotype control antibody and recombinant human IgG1 isotype control antibodies are available. Condition of sample preparation and optimal sample dilution should be determined experimentally by the investigator.
Order Offline	Phone: 1-617-401-8149 Fax: 1-617-606-5022 Email: message@sydlabs.com Or leave a message with a formal purchase order (PO) Or credit card.

Description

PA007274.r2a: Recombinant Anti-mouse CD40 Monoclonal Antibody(Clone: FGK4.5), Rat IgG2a Kappa, In vivo Grade

The anti-mouse CD40 monoclonal antibody (clone: FGK4.5) was produced in mammalian cells.
The FGK4.5 antibody binds to the cell-surface molecule CD40, a member of the tumor necrosis factor receptor superfamily. CD40 is expressed by antigen-presenting cells (APC) and many tumor cells.

References for Recombinant Anti-mouse CD40 Antibody(Clone: FGK4.5):

1、Induction of potent anti-tumor responses while eliminating systemic side effects via liposome-anchored combinatorial immunotherapy
Brandon Kwong,et al.Biomaterials. 2012.PMCID: PMC3140866
“Immunostimulatory therapies that activate immune response pathways are of great interest for overcoming the immunosuppression present in advanced tumors. Agonistic anti-CD40 antibodies and CpG oligonucleotides have previously demonstrated potent, synergistic anti-tumor effects, but their clinical use even as monotherapies is hampered by dose-limiting inflammatory toxicity provoked upon systemic exposure. We hypothesized that by anchoring immuno-agonist compounds to lipid nanoparticles we could retain the bio-activity of therapeutics in the local tumor tissue and tumor-draining lymph node, but limit systemic exposure to these potent molecules. We prepared PEGylated liposomes bearing surface-conjugated anti-CD40 and CpG and assessed their therapeutic efficacy and systemic toxicity compared to soluble versions of the same immuno-agonists, injected intratumorally in the B16F10 murine model of melanoma. Anti-CD40/CpG-liposomes significantly inhibited tumor growth and induced a survival benefit similar to locally injected soluble anti-CD40+CpG. Biodistribution analyses following local delivery showed that the liposomal carriers successfully sequestered anti-CD40 and CpG in vivo, reducing leakage into systemic circulation while allowing draining to the tumor-proximal lymph node. Contrary to locally administered soluble immunotherapy, anti-CD40/CpG liposomes did not elicit significant increases in serum levels of ALT enzyme, systemic inflammatory cytokines, or overall weight loss, confirming that off-target inflammatory effects had been minimized. The development of a delivery strategy capable of inducing robust anti-tumor responses concurrent with minimal systemic side effects is crucial for the continued progress of potent immunotherapies toward widespread clinical translation.”

2、SAP modulates B cell functions in a genetic background-dependent manner
Cynthia Detre,et al.Immunol Lett. 2014.PMCID: PMC3758805
“Mutations affecting the SLAM-associated protein (SAP) are responsible for the X-linked lympho-proliferative syndrome (XLP), a severe primary immunodeficiency syndrome with disease manifestations that include fatal mononucleosis, B cell lymphoma and dysgammaglobulinemia. It is well accepted that insufficient help by SAP−/− CD4+ T cells, in particular during the germinal center reaction, is a component of dysgammaglobulinemia in XLP patients and SAP−/− animals. It is however not well understood whether in XLP patients and SAP−/− mice B cell functions are affected, even though B cells themselves do not express SAP. Here we report that B cell intrinsic responses to haptenated protein antigens are impaired in SAP−/− mice and in Rag−/− mice into which B cells derived from SAP−/− mice together with wt CD4+ T cells had been transferred. This impaired B cells functions are in part depending on the genetic background of the SAP−/− mouse, which affects B cell homeostasis. Surprisingly, stimulation with an agonistic anti-CD40 causes strong in vivo and in vitro B cell responses in SAP−/− mice. Taken together, the data demonstrate that genetic factors play an important role in the SAP-related B cell functions. The finding that anti-CD40 can in part restore impaired B cell responses in SAP−/− mice, suggests potentially novel therapeutic interventions in subsets of XLP patients.”

3、Multimodal intralesional therapy for reshaping the myeloid compartment of tumors resistant to anti-PD-L1 therapy via IRF8
Ankit Patel,et al.J Immunol. 2022.PMCID: PMC8387427
“Intralesional therapy is a promising approach for remodeling the immunosuppressive tumor microenvironment (TME) while minimizing systemic toxicities. A combinatorial in situ immunomodulation (ISIM) regimen with intratumoral administration of Fms-like tyrosine kinase 3 ligand (Flt3L), local radiation, and toll-like receptor 3 (TLR3)/CD40 stimulation induces and activates conventional type 1 dendritic cells (cDC1s) in the TME, and elicits de novo adaptive T cell immunity in poorly T cell-inflamed tumors. However, the impact of ISIM on myeloid-derived suppressor cells (MDSCs) that may promote treatment resistance remains unknown. Here, we examined changes in the frequencies and heterogeneity of CD11b+Ly6CloLy6G+ polymorphonuclear (PMN)- and CD11b+Ly6ChiLy6G− monocytic (M)-MDSCs in ISIM-treated tumors using mouse models of triple-negative breast cancer (TNBC). We found that ISIM treatment decreased intratumoral PMN-MDSCs, but not M-MDSCs. Although the frequency of M-MDSCs remained unchanged, ISIM caused a substantial reduction of CX3CR1+ M-MDSCs that express F4/80. Importantly, these ISIM-induced changes in tumor-residing MDSCs were not observed in Batf3−/− mice. ISIM upregulated PD-L1 expression in both M-MDSCs and PMN-MDSCs, and synergized with anti-PD-L1 therapy. Furthermore, ISIM increased the expression of interferon regulatory factor-8 (IRF8) in myeloid cells, a known negative regulator of MDSCs, indicating a potential mechanism by which ISIM decreases PMN-MDSC levels. Accordingly, ISIM-mediated reduction of PMN-MDSCs was not observed in mice with conditional deletion of IRF8 in myeloid cells. Altogether, these findings suggest that ISIM holds promise as a multimodal intralesional therapy to alter both lymphoid and myeloid compartments of highly aggressive poorly T cell-inflamed, myeloid-enriched tumors resistant to anti-PD-L1 therapy.”

4、Treatment of pancreatic cancer with irreversible electroporation and intratumoral CD40 antibody stimulates systemic immune responses that inhibit liver metastasis in an orthotopic model
Jayanth S Shankara Narayanan,et al.J Immunother Cancer. 2022.PMCID: PMC9843215
“Pancreatic cancer (PC) has a poor prognosis, and most patients present with either locally advanced or distant metastatic disease. Irreversible electroporation (IRE) is a non-thermal method of ablation used clinically in locally advanced PC, but most patients eventually develop distant recurrence. We have previously shown that IRE alone is capable of generating protective, neoantigen-specific immunity. Here, we aim to generate meaningful therapeutic immune effects by combining IRE with local (intratumoral) delivery of a CD40 agonistic antibody (CD40Ab).

Methods
KPC46 organoids were generated from a tumor-bearing male KrasLSL-G12D-p53LSL-R172H-Pdx-1-Cre (KPC) mouse. Orthotopic tumors were established in the pancreatic tail of B6/129 F1J mice via laparotomy. Mice were randomized to treatment with either sham laparotomy, IRE alone, CD40Ab alone, or IRE followed immediately by CD40Ab injection. Metastatic disease and immune infiltration in the liver were analyzed 14 days postprocedure using flow cytometry and multiplex immunofluorescence imaging with spatial analysis. Candidate neoantigens were identified by mutanome profiling of tumor tissue for ex vivo functional analyses.”

5、Innate CD8αα+ lymphocytes enhance anti‐CD40 antibody‐mediated colitis in mice
Aaram A Kumar,et al.Immun Inflamm Dis. 2017.PMCID: PMC5418141
“Immune responses in the intestines require tight regulation to avoid uncontrolled inflammation. We previously described an innate lymphocyte population in the intestinal epithelium (referred to as innate CD8αα+, or iCD8α cells) that can protect against gastrointestinal infections such as those mediated by Citrobacter rodentium.

Methods
Here, we have evaluated the potential contribution of these cells to intestinal inflammation by analyzing inflammation development in mice with decreased numbers of iCD8α cells. We also determined the potential of iCD8α cells to secrete granzymes and their potential role during inflammatory processes.”

6、NFAM1 Promotes Pro-Inflammatory Cytokine Production in Mouse and Human Monocytes
Kathryn W Juchem,et al.Front Immunol. 2022.PMCID: PMC8793151
“NFAT activating protein with ITAM motif 1 (NFAM1) is an ITAM bearing-transmembrane receptor that has been reported to play a role in B cell signaling and development. We performed expression analysis of NFAM1 using publicly available gene expression data sets and found that NFAM1 expression is significantly induced in intestinal biopsies from Crohn’s disease (CD) and ulcerative colitis (UC) patients. At the cellular level, we further observed high expression of NFAM1 in monocytes and neutrophils, and low expression in B and T cells. To explore the role of NFAM1 in multiple immune cells and its potential role in IBD, we generated NFAM1-/- mice. In contrast with previous reports using NFAM1-transgenic mice, NFAM1-/- mice have no obvious defects in immune cell development, or B cell responses. Interestingly, NFAM1-/- monocytes produce reduced levels of TNF-α in response to activation by multiple IBD-relevant stimuli, including CD40L, TLR ligands and MDP. Additional cytokines and chemokines such as IL-6, IL-12, CCL3 and CCL4 are also reduced in CD40L stimulated NFAM1-/- monocytes. Collectively, these findings indicate that NFAM1 promotes monocyte activation, thereby amplifying the response to diverse stimuli. Similarly, we observed that deletion of NFAM1 in human monocytes reduces expression of CD40L-induced CCL4. Lastly, to assess the role of NFAM1 in IBD, we compared development of anti-CD40 induced colitis in NFAM1+/+ and NFAM1-/- mice. We found that although NFAM1 deletion had no impact on development of gut pathology, we did observe a decrease in serum TNF-α, confirming that NFAM1 promotes pro-inflammatory cytokine production in vivo. Taken together, we conclude that NFAM1 functions to amplify cytokine production and should be further evaluated as a therapeutic target for treatment of autoimmune disease.”

7、Agonistic CD40 therapy induces tertiary lymphoid structures but impairs responses to checkpoint blockade in glioma
Luuk van Hooren,et al.Nat Commun. 2021.PMCID: PMC8257767
“Gliomas are brain tumors characterized by an immunosuppressive microenvironment. Immunostimulatory agonistic CD40 antibodies (αCD40) are in clinical development for solid tumors, but are yet to be evaluated for glioma. Here, we demonstrate that systemic delivery of αCD40 in preclinical glioma models induces the formation of tertiary lymphoid structures (TLS) in proximity of meningeal tissue. In treatment-naïve glioma patients, the presence of TLS correlates with increased T cell infiltration. However, systemic delivery of αCD40 induces hypofunctional T cells and impairs the response to immune checkpoint inhibitors in pre-clinical glioma models. This is associated with a systemic induction of suppressive CD11b+ B cells post-αCD40 treatment, which accumulate in the tumor microenvironment. Our work unveils the pleiotropic effects of αCD40 therapy in glioma and reveals that immunotherapies can modulate TLS formation in the brain, opening up for future opportunities to regulate the immune response.”

8、Augmented interleukin (IL) -15Rα expression by CD40 activation is critical in synergistic CD8 T-cell mediated antitumor activity of anti-CD40 antibody with IL-15 in TRAMP-C2 tumors in mice
Meili Zhang,et al.J Immunol. 2012.PMCID: PMC3370156
“Interleukin-15 (IL-15) has potential as an immunotherapeutic agent for cancer treatment because it is a critical factor for the proliferation and activation of natural killer (NK) and CD8+ T cells. However, monotherapy of patients with malignancy with IL-15 that has been initiated may not be optimal due to limited expression of the private receptor, IL-15Rα. We demonstrated greater CD8 T-cell mediated therapeutic efficacy of a combination regimen of murine IL-15 (mIL-15) administered with an agonistic anti-CD40 antibody (FGK4.5) that led to increased IL-15Rα expression on dendritic cells (DCs) as well as other cell types in a syngeneic established TRAMP-C2 tumor model. Seventy-100% of TRAMP-C2 tumor-bearing wild type C57BL/6 mice in the combination group manifested sustained remissions whereas only 0–30% in the anti-CD40 alone group and none in the mIL-15 alone group became tumor free (p<0.001). However, the combination regimen showed less efficacy in TRAMP-C2 tumor-bearing IL-15Rα−/− mice than that in wild type mice. The combination regimen significantly increased the numbers of TRAMP-C2 tumor specific SPAS-1/SNC9-H8 tetramer+CD8+ T-cells which were associated with the protection from tumor development on rechallenge with TRAMP-C2 tumor cells. Using an in vitro cytolytic assay that involved NK cells primed by wild type or IL-15Rα−/− bone marrow derived DCs (BMDCs), we demonstrated that the expression of IL-15Rα by DCs appeared to be required for optimal IL-15 induced NK priming and killing. These findings support the view that anti-CD40 mediated augmented IL-15Rα expression was critical in IL-15 associated sustained remissions observed in TRAMP-C2 tumor-bearing mice receiving combination therapy.”

9、CD4 T cell help via B cells is required for lymphopenia-induced CD8 T cell proliferation
Katayoun Ayasoufi,et al.J Immunol. 2022.PMCID: PMC4772151
“Antibody-mediated lymphoablation is commonly used in solid organ and hematopoietic cell transplantation. However, these strategies fail to efficiently control pathogenic memory T cells and significantly improve long term transplant outcomes. Understanding the mechanisms of T cell reconstitution is critical for enhancing the efficacy of antibody-mediated depletion in sensitized recipients. Using a murine analog of anti-thymocyte globulin (mATG) in a mouse model of cardiac transplantation, we previously showed that peritransplant lymphocyte depletion induces rapid memory T cell proliferation and only modestly prolongs allograft survival. We now report that T cell repertoire following depletion is dominated by memory CD4 T cells. Additional depletion of these residual CD4 T cells severely impairs the recovery of memory CD8 T cells after mATG treatment. The CD4 T cell help during CD8 T cell recovery depends on the presence of B cells expressing CD40 and intact CD40/CD154 interactions. The requirement for CD4 T cell help is not limited to the use of mATG in heart allograft recipients, and is observed in non-transplanted mice and after CD8 T cell depletion with mAb instead of mATG. Most importantly, limiting helper signals increases the efficacy of mATG in controlling memory T cell expansion and significantly extends heart allograft survival in sensitized recipients. Our findings uncover the novel role for helper memory CD4 T cells during homeostatic CD8 T cell proliferation and open new avenues for optimizing lymphoablative therapies in allosensitized patients.”

10、Combination of an agonistic anti-CD40 monoclonal antibody and the COX-2 inhibitor celecoxib induces anti-glioma effects by promotion of type-1 immunity in myeloid cells and T-cells
Akemi Kosaka,et al.Cancer Immunol Immunother. 2014.PMCID: PMC4221287
“Malignant gliomas are heavily infiltrated by immature myeloid cells that mediate immunosuppression. Agonistic CD40 monoclonal antibody (mAb) has been shown to activate myeloid cells and promote antitumor immunity. Our previous study has also demonstrated blockade of cyclooxygenase-2 (COX-2) reduces immunosuppressive myeloid cells, thereby suppressing glioma development in mice. We therefore hypothesized that a combinatory strategy to modulate myeloid cells via two distinct pathways, i.e., CD40/CD40L stimulation and COX-2 blockade, would enhance anti-glioma immunity. We used three different mouse glioma models to evaluate therapeutic effects and underlying mechanisms of a combination regimen with an agonist CD40 mAb and the COX-2 inhibitor celecoxib. Treatment of glioma-bearing mice with the combination therapy significantly prolonged survival compared with either anti-CD40 mAb or celecoxib alone. The combination regimen promoted maturation of CD11b+ cells in both spleen and brain, and enhanced Cxcl10 while suppressing Arg1 in CD11b+Gr-1+ cells in the brain. Anti-glioma activity of the combination regimen was T-cell dependent because depletion of CD4+ and CD8+ cells in vivo abrogated the anti-glioma effects. Furthermore, the combination therapy significantly increased the frequency of CD8+ T-cells, enhanced IFN-γ-production and reduced CD4+CD25+Foxp3+ T regulatory cells in the brain, and induced tumor-antigen-specific T-cell responses in lymph nodes. Our findings suggest that the combination therapy of anti-CD40 mAb with celecoxib enhances anti-glioma activities via promotion of type-1 immunity both in myeloid cells and T-cells.”

11、Interleukin-15 combined with an anti-CD40 antibody provides enhanced therapeutic efficacy for murine models of colon cancer
Meili Zhang,et al.Proc Natl Acad Sci U S A. 2009.PMCID: PMC2678620
“IL-15 has potential as an immunotherapeutic agent for cancer treatment because it is a critical factor for the proliferation and activation of natural killer (NK) and CD8+ T cells. Administration of anti-CD40 antibodies has shown anti-tumor effects in vivo through a variety of mechanisms. Furthermore, activation of CD40 led to increased expression of IL-15 receptor-α by dendritic cells, an action that is critical for trans-presentation of IL-15 to NK and CD8+ T cells. In this study, we investigated the therapeutic efficacy of the combination regimen of murine IL-15 (mIL-15) with an agonistic anti-CD40 antibody (FGK4.5) in murine lung metastasis models involving CT26 and MC38, which are murine colon cancer cell lines syngeneic to BALB/c and C57BL/6 mice, respectively. Treatment with mIL-15 or the anti-CD40 antibody alone significantly prolonged survival of both CT26 and MC38 tumor-bearing mice compared with the mice in the PBS solution control group (P < 0.01). Furthermore, combination therapy with both mIL-15 and the anti-CD40 antibody provided greater therapeutic efficacy as demonstrated by prolonged survival of the mice compared with either mIL-15 or the anti-CD40 antibody-alone groups (P < 0.001). We found that NK cells isolated from the mice that received the combination regimen expressed increased levels of intracellular granzyme B and showed stronger cytotoxic activity on the target cells. The findings from this study provide the scientific basis for clinical trials using the combination regimen of IL-15 with an anti-CD40 antibody for the treatment of patients with cancer.”

12、Activation of 4-1BBL+ B cells with CD40 agonism and IFNγ elicits potent immunity against glioblastoma
Catalina Lee-Chang,et al.J Exp Med. 2020.PMCID: PMC7527974
“Immunotherapy has revolutionized the treatment of many tumors. However, most glioblastoma (GBM) patients have not, so far, benefited from such successes. With the goal of exploring ways to boost anti-GBM immunity, we developed a B cell–based vaccine (BVax) that consists of 4-1BBL+ B cells activated with CD40 agonism and IFNγ stimulation. BVax migrates to key secondary lymphoid organs and is proficient at antigen cross-presentation, which promotes both the survival and the functionality of CD8+ T cells. A combination of radiation, BVax, and PD-L1 blockade conferred tumor eradication in 80% of treated tumor-bearing animals. This treatment elicited immunological memory that prevented the growth of new tumors upon subsequent reinjection in cured mice. GBM patient–derived BVax was successful in activating autologous CD8+ T cells; these T cells showed a strong ability to kill autologous glioma cells. Our study provides an efficient alternative to current immunotherapeutic approaches that can be readily translated to the clinic.”

13、Anti-CD40 Monoclonal Antibody Synergizes with CTLA-4 Ig in Promoting Long-Term Graft Survival in Murine Models of Transplantation1
Christopher R Gilson,et al.J Immunol. 2010.PMCID: PMC2828346
“Blockade of the CD40/CD154 signaling pathway using anti-CD154 antibodies has shown promise in attenuating the alloimmune response and promoting long-term graft survival in murine model systems. Unfortunately, thromboembolic side effects observed in humans have hampered its progression through clinical trials. Appropriately designed anti-CD40 antibodies may provide a suitable alternative. We investigated two isoforms of a novel monoclonal rat anti-mouse CD40 antibody (7E1) for characteristics and effects mirroring those of anti-CD154: 7E1-G1 (an IgG1 isotype) and 7E1-G2b (an IgG2b isotype). In vitro proliferation assays to measure the agonist properties of the two anti-CD40 antibodies revealed similar responses when plate-bound. However, when present as a soluble stimulus, 7E1-G1 but not 7E1-G2b led to proliferation. Importantly, 7E1-G2b was as effective as anti-CD154 when administered in vivo in concert with CTLA4-Ig in promoting both allogeneic bone marrow chimerism and skin graft survival, while 7E1-G1 was not. The protection observed with 7E1-G2b was not due to depletion of CD40 bearing antigen presenting cells. These data suggest that an appropriately designed anti-CD40 antibody can promote graft survival as well as anti-CD154, making 7E1-G2b an attractive substitute in mouse models of costimulation blockade-based tolerance regimens.”

14、LUBAC Suppresses IL-21-Induced Apoptosis in CD40-Activated Murine B Cells and Promotes Germinal Center B Cell Survival and the T-Dependent Antibody Response
Jingwei Wang,et al.Front Immunol. 2021.PMCID: PMC8089397
“B cell activation by Tfh cells, i.e., through CD154 engagement of CD40 and IL-21, and survival within GCs are crucial for the T-dependent Ab response. LUBAC, composed of HOIP, SHARPIN, and HOIL-1, catalyzes linear ubiquitination (Linear M1-Ub) to mediate NF-κB activation and cell survival induced by TNF receptor superfamily members, which include CD40. As shown in this study, B cells expressing the Sharpin null mutation cpdm (Sharpincpdm) could undergo proliferation, CSR, and SHM in response to immunization by a T-dependent Ag, but were defective in survival within GCs, enrichment of a mutation enhancing the BCR affinity, and production of specific Abs. Sharpincpdm B cells stimulated in vitro with CD154 displayed normal proliferation and differentiation, marginally impaired NF-κB activation and survival, but markedly exacerbated death triggered by IL-21. While activating the mitochondria-dependent apoptosis pathway in both Sharpin+/+ and Sharpincpdm B cells, IL-21 induced Sharpincpdm B cells to undergo sustained activation of caspase 9 and caspase 8 of the mitochondria-dependent and independent pathway, respectively, and ultimately caspase 3 in effecting apoptosis. These were associated with loss of the caspase 8 inhibitor cFLIP and reduction in cFLIP Linear M1-Ub, which interferes with cFLIP poly-ubiquitination at Lys48 and degradation. Finally, the viability of Sharpincpdm B cells was rescued by caspase inhibitors but virtually abrogated – together with Linear M1-Ub and cFLIP levels – by a small molecule HOIP inhibitor. Thus, LUBAC controls the cFLIP expression and inhibits the effects of caspase 8 and IL-21-activated caspase 9, thereby suppressing apoptosis of CD40 and IL-21-activated B cells and promoting GC B cell survival.”

15、B cells orchestrate tolerance to the neuromyelitis optica autoantigen AQP4
Ali Maisam Afzali,et al.Nature. 2024.PMCID: PMC10937377
“Neuromyelitis optica is a paradigmatic autoimmune disease of the central nervous system, in which the water-channel protein AQP4 is the target antigen1. The immunopathology in neuromyelitis optica is largely driven by autoantibodies to AQP42. However, the T cell response that is required for the generation of these anti-AQP4 antibodies is not well understood. Here we show that B cells endogenously express AQP4 in response to activation with anti-CD40 and IL-21 and are able to present their endogenous AQP4 to T cells with an AQP4-specific T cell receptor (TCR). A population of thymic B cells emulates a CD40-stimulated B cell transcriptome, including AQP4 (in mice and humans), and efficiently purges the thymic TCR repertoire of AQP4-reactive clones. Genetic ablation of Aqp4 in B cells rescues AQP4-specific TCRs despite sufficient expression of AQP4 in medullary thymic epithelial cells, and B-cell-conditional AQP4-deficient mice are fully competent to raise AQP4-specific antibodies in productive germinal-centre responses. Thus, the negative selection of AQP4-specific thymocytes is dependent on the expression and presentation of AQP4 by thymic B cells. As AQP4 is expressed in B cells in a CD40-dependent (but not AIRE-dependent) manner, we propose that thymic B cells might tolerize against a group of germinal-centre-associated antigens, including disease-relevant autoantigens such as AQP4.”

16、Nanomaterials-Mediated Co-Stimulation of Toll-Like Receptors and CD40 for Antitumor Immunity
Jingyue Yan,et al.Adv Mater. 2023.PMCID: PMC9691606
“Toll-like receptors (TLRs) and CD40-related signaling pathways represent critical bridges between innate and adaptive immune responses. Here, an immunotherapy regimen that enables co-stimulation of TLR7/8- and CD40-mediated pathways is developed. TLR7/8 agonist resiquimod (R848) derived amino lipids, RAL1 and RAL2, are synthesized and formulated into RAL-derived lipid nanoparticles (RAL-LNPs). The RAL2-LNPs show efficient CD40 mRNA delivery to DCs both in vitro (90.8 ± 2.7%) and in vivo (61.3 ± 16.4%). When combined with agonistic anti-CD40 antibody, this approach can produce effective antitumor activities in mouse melanoma tumor models, thereby suppressing tumor growth, prolonging mouse survival, and establishing antitumor memory immunity. Overall, RAL2-LNPs provide a novel platform toward cancer immunotherapy by integrating innate and adaptive immunity.”

17、The quantity of CD40 signaling determines the differentiation of B cells into functionally distinct memory cell subsets
Takuya Koike,et al.eLife. 2019.PMCID: PMC6636905
“In mice, memory B (Bmem) cells can be divided into two subpopulations: CD80hi Bmem cells, which preferentially differentiate into plasma cells; and CD80lo Bmem cells, which become germinal center (GC) B cells during a recall response. We demonstrate that these distinct responses can be B-cell-intrinsic and essentially independent of B-cell receptor (BCR) isotypes. Furthermore, we find that the development of CD80hi Bmem cells in the primary immune response requires follicular helper T cells, a relatively strong CD40 signal and a high-affinity BCR on B cells, whereas the development of CD80lo Bmem cells does not. Quantitative differences in CD40 stimulation were enough to recapitulate the distinct B cell fate decisions in an in vitro culture system. The quantity of CD40 signaling appears to be translated into NF-κB activation, followed by BATF upregulation that promotes Bmem cell differentiation from GC B cells..”

18、Macrophages and CD8+ T cells mediate the antitumor efficacy of combined CD40 ligation and imatinib therapy in gastrointestinal stromal tumors
Jennifer Q Zhang,et al.Cancer Immunol Res. 2018.PMCID: PMC6203303
“Tyrosine kinase inhibition of gastrointestinal stromal tumors (GIST) is effective but typically culminates in resistance and is rarely curative. Immunotherapy has potential application to GIST, as we previously showed that T-cell checkpoint blockade increases the antitumor effects of imatinib. Here, we showed that ligation of CD40 using an agonistic antibody (anti-CD40) activated tumor-associated macrophages (TAMs) in vivo in a knock-in mouse model of GIST harboring a germline mutation in Kit exon 11. Activated TAMs had greater TNF production and NFκB signaling and directly inhibited tumor cells in vitro. Anti-CD40 required concomitant therapy with imatinib for efficacy and depended on TAMs, and to a lesser extent CD8+ T cells, but not on CD4+ T cells or B cells. In an analysis of 50 human GIST specimens by flow cytometry, we found that CD40 was expressed on human TAMs and tumor cells yet was downregulated after response to imatinib. CD40 ligation did not have a direct inhibitory effect on human GIST cells. Our findings provide the rationale for combining anti-CD40 and tyrosine kinase inhibition to treat human GIST.”

19、Mechanisms of CD40-dependent cDC1 licensing beyond costimulation
Renee Wu,et al.Nat Immunol. 2023.PMCID: PMC9896965
“CD40 signaling in classical type 1 dendritic cells (cDC1s) is required for CD8 T cell-mediated tumor rejection, but the underlying mechanisms are incompletely understood. Here, we identified CD40-induced genes in cDC1s, including Cd70, Tnfsf9, Ptgs2 and Bcl2l1, and examined their contributions to anti-tumor immunity. cDC1-specific inactivation of CD70 and COX-2, and global CD27 inactivation, only partially impaired tumor rejection or tumor-specific CD8 T cell expansion. Loss of 4-1BB, alone or in Cd27−/− mice, did not further impair anti-tumor immunity. However, cDC1-specific CD40 inactivation reduced cDC1 mitochondrial transmembrane potential and increased caspase activation in tumor-draining lymph nodes, reducing migratory cDC1 numbers in vivo. Similar impairments occurred during in vitro antigen presentation by Cd40−/− cDC1s to CD8+ T cells, which were reversed by re-expression of Bcl2l1. Thus, CD40 signaling in cDC1s not only induces costimulatory ligands for CD8+ T cells but also induces Bcl2l1 that sustains cDC1 survival during priming of anti-tumor responses.”

20、CD40 signaling restricts RNA virus replication in macrophages, leading to rapid innate immune control of acute virus infection
Kai J Rogers,et al.J Leukoc Biol. 2021.PMCID: PMC7774454
“Many acute viral infections target tissue macrophages, yet the mechanisms of macrophage-mediated control of viruses are poorly understood. Here, we report that CD40 expressed by peritoneal macrophages restricts early infection of a broad range of RNA viruses. Loss of CD40 expression enhanced virus replication as early as 12–24 hours of infection and, conversely, stimulation of CD40 signaling with an agonistic antibody blocked infection. With peritoneal cell populations infected with the filovirus, wild-type (WT) Ebola virus (EBOV), or a BSL2 model virus, recombinant vesicular stomatitis virus encoding Ebola virus glycoprotein (rVSV/EBOV GP), we examined the mechanism conferring protection. Here we demonstrate that restricted virus replication in macrophages required CD154/CD40 interactions which stimulated IL-12 production through TRAF6-dependent signaling. In turn, IL-12 production resulted in interferon gamma (IFN-γ) production, which induced proinflammatory polarization of macrophages, protecting the cells from infection. These CD40-dependent events protected mice against virus challenge. CD40−/− mice were exquisitely sensitive to intraperitoneal challenge with a dose of rVSV/EBOV GP that was sublethal to CD40+/+ mice, exhibiting viremia within 12 hours of infection and rapidly succumbing to infection. This study identifies a previously unappreciated role for macrophage-intrinsic CD40 signaling in controlling acute virus infection.”

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Please remember our product information: In vivo Grade Recombinant Anti-mouse CD40 Monoclonal Antibody, Rat IgG2a Kappa (Clone: FGK4.5): PA007274.r2a Syd Labs

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