RESUMEN

OBJECTIVES: Chronic rejection remains the leading cause of progressive decline in graft function. Accumulating evidence indicates that macrophages participate in chronic rejection dependent on CD40-CD40L. The FOS family members are critical in inflammatory and immune responses. However, the mechanisms underlying the role of FOS family members in chronic rejection remain unclear. In this study, we aimed to elucidate the role and underlying mechanisms of FOS-positive macrophages regulated by CD40 that mediate chronic allograft rejection. MATERIALS AND METHODS: We downloaded publicly accessible chronic rejection kidney transplant single-cell sequencing datasets from the gene expression omnibus database. Differentially expressed genes between the CD40hi and CD40low macrophage chronic rejection groups were analyzed. We established a chronic rejection mouse model by using CTLA-4-Ig. We treated bone marrow-derived macrophages with an anti-CD40 antibody. We assessed expression of the FOS family by flow cytometry, real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. We identified altered signaling pathways by using RNA sequencing analysis. We detected DNA specifically bound to transcription factors by using ChIP-sequencing, with detection of the degree of graft fibrosis and survival. RESULTS: FOS was highly expressed on CD40hi macrophages in patients with chronic transplantrejection. Mechanistically, we showed that CD40 activated NF-κB2 translocation into the nucleus to upregulate c-Fos and FosB expression, thus promoting chronic rejection of cardiac transplant.We showed thatNF-κB2 regulated c-Fos and FosB expression by binding to the c-fos and fosb promoter regions. Inhibition of c-Fos/activator protein-1 decreased graft fibrosis and prolonged graft survival. CONCLUSIONS: CD40 may activate transcription factor NF-κB2 translocation into the nucleus of macrophages to upregulate c-Fos and FosB expression, thus promoting chronic rejection of cardiac transplant. Inhibition of c-Fos/activator protein-1 decreased grafts fibrosis and prolonged graft survival.

Asunto(s)

Antígenos CD40 , Modelos Animales de Enfermedad , Rechazo de Injerto , Trasplante de Corazón , Macrófagos , Proteínas Proto-Oncogénicas c-fos , Transducción de Señal , Animales , Humanos , Masculino , Ratones , Antígenos CD40/metabolismo , Antígenos CD40/genética , Células Cultivadas , Enfermedad Crónica , Bases de Datos Genéticas , Fibrosis , Rechazo de Injerto/inmunología , Rechazo de Injerto/metabolismo , Rechazo de Injerto/genética , Supervivencia de Injerto , Trasplante de Corazón/efectos adversos , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , FN-kappa B/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-fos/genética , Factor de Transcripción AP-1/metabolismo

RESUMEN

Glioblastoma multiforme (GBM) is one of the most aggressive and deadly forms of brain cancer, which has a very complex tumor microenvironment (TME) promoting tumor growth, immune evasion, and resistance to therapy. The main players within this environment are represented by cytokines such as Interleukin-4, Interleukin-6, and Interleukin-13, along with the costimulatory molecule CD40. The paper draws back the curtain on the complex interactions played out by these molecules in contributing to the formation of a TME within GBM. IL-4 and IL-13 induce an immunosuppressive environment through the polarization of tumor-associated macrophages (TAMs) into a pro-tumoral M2 phenotype. In contrast, IL-6 takes part in the activation of the JAK-STAT3 pathway, enhancing survival and proliferation of tumor cells. In this context, CD40 either induces anti-tumor immunity through APC activation or facilitates tumors by angiogenesis and survival pathways. The synergistic actions of these molecules create feedback loops that keep up the malignancy of GBM and present a big problem for therapy. Knowledge of these interactions opens new ways for the development of multi-targeted therapeutic strategies at the other end. This may result in the interruption of the tumor-supportive environment in GBM, reducing tumor growth and improving patient outcomes by targeting IL-4, IL-6, IL-13, and CD40 simultaneously.

Asunto(s)

Neoplasias Encefálicas , Antígenos CD40 , Glioblastoma , Interleucina-13 , Interleucina-4 , Interleucina-6 , Microambiente Tumoral , Humanos , Antígenos CD40/metabolismo , Ensayos Clínicos como Asunto , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Interleucina-6/metabolismo

RESUMEN

OBJECTIVE: To observe the effects and mechanisms of Maresin2 on the function of DCs(Dendritic cells). METHOD: The levels of IL-6, IL-12, TNF-α and IL-1ß secreted by BMDCs (Bone marrow-derived Dendritic cells) after Maresin2 treatment were detected by ELISA. At the same time, the expressions of costimulatory molecules CD40 and CD86 on the surface, the ability of phagocytosis of ovalbumin(OVA) antigen, and antigen presentation function in BMDCs were analyzed by flow cytometry. Finally, MAPK and NF-κB pathway signaling phosphorylation in Maresin2-treated BMDCs were detected by western blot. RESULTS: The secretion levels of IL-6, IL-12, TNF-α and IL-1ß were significantly decreased in the Maresin2 treatment group after LPS treatment (P < 0.05). The expression levels of CD86 and CD40 were significantly decreased after Maresin2 treatment (P < 0.05). Maresin2 enhanced the phagocytosis ability of ovalbumin(OVA) (P < 0.05), but the ability of antigen presentation of BMDCs with the treatment of Maresin2 changed slightly (P > 0.05). Phosphorylation of p38, JNK, p65, ikka/ß and ERK peaked at 15 min in the LPS group, while phosphorylation of p-p38 and p-ERK weakened 30 min and 60 min after treatment with Maresin2. CONCLUSIONS: Maresin2 inhibits inflammatory cytokine secretion but enhances phagocytosis via the MAPK/NF-κB pathway in BMDCs, which may contribute to negatively regulating inflammation.

Asunto(s)

Citocinas , Células Dendríticas , FN-kappa B , Fagocitosis , Transducción de Señal , Animales , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , FN-kappa B/metabolismo , Ratones , Citocinas/metabolismo , Transducción de Señal/efectos de los fármacos , Fagocitosis/efectos de los fármacos , Células Cultivadas , Ovalbúmina/inmunología , Lipopolisacáridos/farmacología , Lipopolisacáridos/inmunología , Ratones Endogámicos C57BL , Diferenciación Celular/efectos de los fármacos , Antígenos CD40/metabolismo , Presentación de Antígeno/efectos de los fármacos , Ácidos Docosahexaenoicos

RESUMEN

Visceral leishmaniasis (VL) is a neglected tropical disease caused by parasites from the Leishmania (L.) donovani complex. VL is characterised by uncontrolled parasite replication in spleen, liver and bone marrow, and by an impaired immune response and high systemic levels of inflammation. Monocytes have been poorly characterised in VL patients. The aim of this study was to evaluate the expression levels of markers involved in the regulation of T cell responses on different subsets of monocytes from the blood of VL patients and healthy non-endemic controls (HNEC). Monocytes can broadly be divided into three subsets: classical, intermediate and non-classical monocytes. Our results show that the percentages of all three subsets stayed similar at the time of VL diagnosis (ToD) and at the end of anti-leishmanial treatment (EoT). We first looked at co-stimulatory receptors: the expression levels of CD40 were significantly increased on classical and intermediate, but not non-classical monocytes, at ToD as compared to EoT and HNEC. CD80 expression levels were also increased on intermediate monocytes at ToD as compared to EoT and HNEC, and on classical monocytes only as compared to HNEC. The levels of CD86 were similar at EoT and ToD and in HNEC on classical and intermediate monocytes, but significantly higher at EoT on non-classical monocytes. We also looked at an inhibitory molecule, PD-L1. Our results show that the expression levels of PD-L1 were significantly higher on all three monocyte subsets at ToD as compared to HNEC, and to EoT on classical and intermediate monocytes. These results show that monocytes from the blood of VL patients upregulate both co-stimulatory and inhibitory receptors and that their expression levels are restored at EoT.

Asunto(s)

Antígenos CD40 , Leishmaniasis Visceral , Monocitos , Humanos , Leishmaniasis Visceral/inmunología , Leishmaniasis Visceral/parasitología , Leishmaniasis Visceral/sangre , Monocitos/inmunología , Masculino , Femenino , Adulto , Antígenos CD40/metabolismo , Adulto Joven , Persona de Mediana Edad , Antígeno B7-1/metabolismo , Adolescente , Antígeno B7-2/metabolismo , Leishmania donovani/inmunología

RESUMEN

The combination of radiotherapy/chemoradiotherapy and immune checkpoint blockade can result in poor outcomes in patients with locally advanced head and neck squamous cell carcinoma (HNSCC). Here, we show that combining ATR inhibition (ATRi) with radiotherapy (RT) increases the frequency of activated NKG2A+PD-1+ T cells in animal models of HNSCC. Compared with the ATRi/RT treatment regimen alone, the addition of simultaneous NKG2A and PD-L1 blockade to ATRi/RT, in the adjuvant, post-radiotherapy setting induces a robust antitumour response driven by higher infiltration and activation of cytotoxic T cells in the tumour microenvironment. The efficacy of this combination relies on CD40/CD40L costimulation and infiltration of activated, proliferating memory CD8+ and CD4+ T cells with persistent or new T cell receptor (TCR) signalling, respectively. We also observe increased richness in the TCR repertoire and emergence of numerous and large TCR clonotypes that cluster based on antigen specificity in response to NKG2A/PD-L1/ATRi/RT. Collectively, our data point towards potential combination approaches for the treatment of HNSCC.

Asunto(s)

Proteínas de la Ataxia Telangiectasia Mutada , Antígeno B7-H1 , Inmunoterapia , Carcinoma de Células Escamosas de Cabeza y Cuello , Microambiente Tumoral , Animales , Femenino , Humanos , Ratones , Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Proteínas de la Ataxia Telangiectasia Mutada/antagonistas & inhibidores , Antígeno B7-H1/antagonistas & inhibidores , Antígeno B7-H1/metabolismo , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/efectos de la radiación , Antígenos CD40/metabolismo , Antígenos CD40/inmunología , Antígenos CD40/antagonistas & inhibidores , Linfocitos T CD8-positivos/inmunología , Línea Celular Tumoral , Neoplasias de Cabeza y Cuello/inmunología , Neoplasias de Cabeza y Cuello/terapia , Neoplasias de Cabeza y Cuello/radioterapia , Inhibidores de Puntos de Control Inmunológico/farmacología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inmunoterapia/métodos , Ratones Endogámicos C57BL , Subfamília C de Receptores Similares a Lectina de Células NK/metabolismo , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Receptor de Muerte Celular Programada 1/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/inmunología , Carcinoma de Células Escamosas de Cabeza y Cuello/radioterapia , Carcinoma de Células Escamosas de Cabeza y Cuello/terapia , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Linfocitos T Citotóxicos/inmunología , Microambiente Tumoral/efectos de la radiación

RESUMEN

The involvement of Group 3 innate lymphoid cells (ILC3s) and dendritic cells (DCs) in chronic lung inflammation has been increasingly regarded as the key to understand the inflammatory mechanisms of smoke-related chronic obstructive pulmonary disease (COPD). However, the mechanism underlying the engagement of both remains unclear. Our study aimed to explore NCR-ILC3 differentiation in the lungs of mice exposed to cigarette smoke (CS) and to further investigate whether DCs activated by CS exposure contribute to the differentiation of ILCs into NCR-ILC3s. The study involved both in vivo and in vitro experiments. In the former, the frequencies of lung NCR-ILC3s and NKp46-IL-17A+ ILCs and the expression of DCs, CD40, CD86, IL-23, and IL-1ß quantified by flow cytometry were compared between CS-exposed mice and air-exposed mice. In the latter, NKp46-IL-17A+ ILC frequencies quantified by flow cytometry were compared after two cocultures, one involving lung CD45+Lin-CD127+ ILCs sorted from air-exposed mice and DCs sifted by CD11c magnetic beads from CS-exposed mice and another including identical CD45+Lin-CD127+ ILCs and DCs from air-exposed mice. The results indicated significant increases in the frequencies of NCR-ILC3s and NKp46-IL-17A+ ILCs; in the expression of DCs, CD40, CD86, IL-23, and IL-1ß in CS-exposed mice; and in the frequency of NKp46-IL-17A+ ILCs after the coculture with DCs from CS-exposed mice. In conclusion, CS exposure increases the frequency of lung ILCs and NCR-ILC3s. CS-induced DC activation enhances the differentiation of ILCs into NCR-ILC3s, which likely acts as a mediating step in the involvement of NCR-ILC3s in chronic lung inflammation.

Asunto(s)

Diferenciación Celular , Células Dendríticas , Interleucina-17 , Interleucina-1beta , Pulmón , Receptor 1 Gatillante de la Citotoxidad Natural , Animales , Células Dendríticas/inmunología , Ratones , Pulmón/inmunología , Pulmón/metabolismo , Receptor 1 Gatillante de la Citotoxidad Natural/metabolismo , Interleucina-17/metabolismo , Interleucina-1beta/metabolismo , Linfocitos/inmunología , Linfocitos/metabolismo , Interleucina-23/metabolismo , Antígeno B7-2/metabolismo , Ratones Endogámicos C57BL , Humo/efectos adversos , Enfermedad Pulmonar Obstructiva Crónica/inmunología , Antígenos CD40/metabolismo , Fumar Cigarrillos/efectos adversos , Inmunidad Innata , Antígenos Ly/metabolismo , Técnicas de Cocultivo , Masculino

RESUMEN

Ligation of the B cell antigen receptor (BCR) initiates humoral immunity. However, BCR signaling without appropriate co-stimulation commits B cells to death rather than to differentiation into immune effector cells. How BCR activation depletes potentially autoreactive B cells while simultaneously primes for receiving rescue and differentiation signals from cognate T lymphocytes remains unknown. Here, we use a mass spectrometry-based proteomic approach to identify cytosolic/nuclear shuttling elements and uncover transcription factor EB (TFEB) as a central BCR-controlled rheostat that drives activation-induced apoptosis, and concurrently promotes the reception of co-stimulatory rescue signals by supporting B cell migration and antigen presentation. CD40 co-stimulation prevents TFEB-driven cell death, while enhancing and prolonging TFEB's nuclear residency, which hallmarks antigenic experience also of memory B cells. In mice, TFEB shapes the transcriptional landscape of germinal center B cells. Within the germinal center, TFEB facilitates the dark zone entry of light-zone-residing centrocytes through regulation of chemokine receptors and, by balancing the expression of Bcl-2/BH3-only family members, integrates antigen-induced apoptosis with T cell-provided CD40 survival signals. Thus, TFEB reprograms antigen-primed germinal center B cells for cell fate decisions.

Asunto(s)

Apoptosis , Linfocitos B , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice , Antígenos CD40 , Centro Germinal , Receptores de Antígenos de Linfocitos B , Animales , Centro Germinal/inmunología , Centro Germinal/citología , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/metabolismo , Linfocitos B/inmunología , Linfocitos B/metabolismo , Ratones , Antígenos CD40/metabolismo , Antígenos CD40/inmunología , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores de Antígenos de Linfocitos B/inmunología , Ratones Endogámicos C57BL , Activación de Linfocitos/inmunología , Diferenciación Celular/inmunología , Transducción de Señal , Presentación de Antígeno/inmunología

RESUMEN

Hypertension causes platelet activation and adhesion in the brain resulting in glial activation and neuroinflammation. Further, activation of Angiotensin-Converting Enzyme 2/Angiotensin (1-7)/Mas Receptor (ACE2/Ang (1-7)/MasR) axis of central Renin-Angiotensin System (RAS), is known to reduce glial activation and neuroinflammation, thereby exhibiting anti-hypertensive and anti-neuroinflammatory properties. Therefore, in the present study, the role of ACE2/Ang (1-7)/MasR axis was studied on platelet-induced glial activation and neuroinflammation using Diminazene Aceturate (DIZE), an ACE2 activator, in astrocytes and microglial cells as well as in rat model of hypertension. We found that the ACE2 activator DIZE, independently of its BP-lowering properties, efficiently prevented hypertension-induced glial activation, neuroinflammation, and platelet CD40-CD40L signaling via upregulation of ACE2/Ang (1-7)/MasR axis. Further, DIZE decreased platelet deposition in the brain by reducing the expression of adhesion molecules on the brain endothelium. Activation of ACE2 also reduced hypertension-induced endothelial dysfunction by increasing eNOS bioavailability. Interestingly, platelets isolated from hypertensive rats or activated with ADP had significantly increased sCD40L levels and induced significantly more glial activation than platelets from DIZE treated group. Therefore, injection of DIZE pre-treated ADP-activated platelets into normotensive rats strongly reduced glial activation compared to ADP-treated platelets. Moreover, CD40L-induced glial activation, CD40 expression, and NFкB-NLRP3 inflammatory signaling are reversed by DIZE. Furthermore, the beneficial effects of ACE2 activation, DIZE was found to be significantly blocked by MLN4760 (ACE2 inhibitor) as well as A779 (MasR antagonist) treatments. Hence, our study demonstrated that ACE2 activation reduced the platelet CD40-CD40L induced glial activation and neuroinflammation, hence imparted neuroprotection.

Asunto(s)

Enzima Convertidora de Angiotensina 2 , Ligando de CD40 , Diminazeno , Modelos Animales de Enfermedad , Hipertensión , Peptidil-Dipeptidasa A , Transducción de Señal , Animales , Diminazeno/análogos & derivados , Diminazeno/farmacología , Diminazeno/uso terapéutico , Enzima Convertidora de Angiotensina 2/metabolismo , Masculino , Transducción de Señal/efectos de los fármacos , Hipertensión/tratamiento farmacológico , Ligando de CD40/metabolismo , Peptidil-Dipeptidasa A/metabolismo , Ratas , Plaquetas/efectos de los fármacos , Plaquetas/metabolismo , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Proto-Oncogenes Mas , Neuroglía/efectos de los fármacos , Neuroglía/metabolismo , Fragmentos de Péptidos , Angiotensina I , Células Cultivadas , Microglía/efectos de los fármacos , Microglía/metabolismo , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Ratas Wistar , Sistema Renina-Angiotensina/efectos de los fármacos , Receptores Acoplados a Proteínas G/metabolismo , Antígenos CD40/metabolismo , Humanos , Activación Plaquetaria/efectos de los fármacos , Antihipertensivos/farmacología , Antihipertensivos/uso terapéutico

RESUMEN

BACKGROUND: Heterogeneity of cerebral atrophic rate commonly exists in mild cognitive impairment (MCI), which may be associated with microglia-involved neuropathology and have an influence on cognitive outcomes. OBJECTIVE: We aim to explore the heterogeneity of cerebral atrophic rate among MCI and its association with plasma proteins related to microglia activity, with further investigation of their interaction effects on long-term cognition. SUBJECTS: A total of 630 MCI subjects in the ADNI database were included, of which 260 subjects were available with baseline data on plasma proteins. METHODS: Group-based multi-trajectory modeling (GBMT) was used to identify the latent classes with heterogeneous cerebral atrophic rates. Associations between latent classes and plasma proteins related to microglia activity were investigated with generalized linear models. Linear mixed effect models (LME) were implemented to explore the interaction effects between proteins related to microglia activity and identified latent classes on longitudinal cognitive changes. RESULTS: Two latent classes were identified and labeled as the slow-atrophy class and the fast-atrophy class. Associations were found between such heterogeneity of atrophic rates and plasma proteins related to microglia activity, especially AXL receptor tyrosine kinase (AXL), CD40 antigen (CD40), and tumor necrosis factor receptor-like 2 (TNF-R2). Interaction effects on longitudinal cognitive changes showed that higher CD40 was associated with faster cognitive decline in the slow-atrophy class and higher AXL or TNF-R2 was associated with slower cognitive decline in the fast-atrophy class. CONCLUSIONS: Heterogeneity of atrophic rates at the MCI stage is associated with several plasma proteins related to microglia activity, which show either protective or adverse effects on long-term cognition depending on the variability of atrophic rates.

Asunto(s)

Atrofia , Disfunción Cognitiva , Microglía , Humanos , Microglía/patología , Masculino , Femenino , Anciano , Atrofia/patología , Estudios Longitudinales , Cognición/fisiología , Anciano de 80 o más Años , Progresión de la Enfermedad , Antígenos CD40/sangre , Proteínas Tirosina Quinasas Receptoras , Receptores Tipo II del Factor de Necrosis Tumoral/sangre , Imagen por Resonancia Magnética , Proteínas Proto-Oncogénicas/sangre , Proteínas Sanguíneas/análisis

RESUMEN

CD40 agonist antibodies (αCD40) have shown promising anti-tumor response in both preclinical and early clinical studies. However, its systemic administration is associated with immune- and hepato-toxicities which hampers its clinical usage. In addition, αCD40 showed low tumor retention and induced PD-L1 expression which makes tumor microenvironment (TME) immunosuppressive. To overcome these issues, in this study, we have developed a multifunctional Immunosome where αCD40 is conjugated on the surface and RRX-001, a small molecule immunomodulator was encapsulated inside it. Immunosomes showed higher tumor accumulation till 96 h of administration and displayed sustained release of αCD40 in vivo. Immunosomes significantly delayed tumor growth and showed tumor free survival in mice bearing GL-261 glioblastoma by increasing the population of CD45+CD8+ T cells, CD45+CD20+ B cells, CD45+CD11c+ DCs and F4/80+CD86+ cells in TME. Immunosome significantly reduced the population of T-regulatory cells, M2 macrophage, and MDSCs and lowered the PD-L1 expression. Moreover, Immunosomes significantly enhanced the levels of Th1 cytokines (IFN-γ, IL-6, IL-2) over Th2 cytokines (IL-4 and IL-10) which supported anti-tumor response. Most interestingly, Immunosomes averted the in vivo toxicities associated with free αCD40 by lowering the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), IL-6, IL-1α and reduced the degree of liver damage. In addition, Immunosomes treated long-term surviving mice showed tumor specific immune memory response which prevented tumor growth upon rechallenge. Our results suggested that this novel formulation can be further explored in clinics to improve in vivo anti-tumor efficacy of αCD40 with long-lasting tumor specific immunity while reducing the associated toxicities.

Asunto(s)

Antígenos CD40 , Glioblastoma , Microambiente Tumoral , Animales , Microambiente Tumoral/efectos de los fármacos , Glioblastoma/tratamiento farmacológico , Glioblastoma/patología , Glioblastoma/inmunología , Antígenos CD40/metabolismo , Ratones , Línea Celular Tumoral , Ratones Endogámicos C57BL , Femenino , Citocinas/metabolismo , Humanos

RESUMEN

Expression of the transcriptional regulator ZFP318 is induced in germinal center (GC)-exiting memory B cell precursors and memory B cells (MBCs). Using a conditional ZFP318 fluorescence reporter that also enables ablation of ZFP318-expressing cells, we found that ZFP318-expressing MBCs were highly enriched with GC-derived cells. Although ZFP318-expressing MBCs constituted only a minority of the antigen-specific MBC compartment, their ablation severely impaired recall responses. Deletion of Zfp318 did not alter the magnitude of primary responses but markedly reduced MBC participation in recall. CD40 ligation promoted Zfp318 expression, whereas B cell receptor (BCR) signaling was inhibitory. Enforced ZFP318 expression enhanced recall performance of MBCs that otherwise responded poorly. ZFP318-deficient MBCs expressed less mitochondrial genes, had structurally compromised mitochondria, and were susceptible to reactivation-induced cell death. The abundance of ZFP318-expressing MBCs, instead of the number of antigen-specific MBCs, correlated with the potency of prime-boost vaccination. Therefore, ZFP318 controls the MBC recallability and represents a quality checkpoint of humoral immune memory.

Asunto(s)

Centro Germinal , Memoria Inmunológica , Células B de Memoria , Mitocondrias , Animales , Mitocondrias/metabolismo , Mitocondrias/inmunología , Ratones , Memoria Inmunológica/genética , Memoria Inmunológica/inmunología , Células B de Memoria/inmunología , Células B de Memoria/metabolismo , Centro Germinal/inmunología , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores de Antígenos de Linfocitos B/genética , Regulación de la Expresión Génica , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Ratones Endogámicos C57BL , Ratones Noqueados , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Transducción de Señal/inmunología , Antígenos CD40/metabolismo , Antígenos CD40/genética , Antígenos CD40/inmunología , Inmunidad Humoral , Transcripción Genética , Proteínas de la Membrana , Proteínas Mitocondriales

RESUMEN

In situ vaccines (ISVs) utilize the localized delivery of chemotherapeutic agents or radiotherapy to stimulate the release of endogenous antigens from tumors, thereby eliciting systemic and persistent immune activation. Recently, a bioinspired ISV strategy has attracted tremendous attention due to its features such as an immune adjuvant effect and genetic plasticity. M13 bacteriophages are natural nanomaterials with intrinsic immunogenicity, genetic flexibility, and cost-effectiveness for large-scale production, demonstrating the potential for application in cancer vaccines. In this study, we propose an ISV based on the engineered M13 bacteriophage targeting CD40 (M13CD40) for dendritic cell (DC)-targeted immune stimulation, named H-GM-M13CD40. We induce immunogenic cell death and release tumor antigens through local delivery of (S)-10-hydroxycamptothecin (HCPT), followed by intratumoral injection of granulocyte-macrophage colony stimulating factor (GM-CSF) and M13CD40 to enhance DC recruitment and activation. We demonstrate that this ISV strategy can result in significant accumulation and activation of DCs at the tumor site, reversing the immunosuppressive tumor microenvironment. In addition, H-GM-M13CD40 can synergize with the PD-1 blockade and induce abscopal effects in cold tumor models. Overall, our study verifies the immunogenicity of the engineered M13CD40 bacteriophage and provides a proof of concept that the engineered M13CD40 phage can function as an adjuvant for ISVs.

Asunto(s)

Bacteriófago M13 , Vacunas contra el Cáncer , Células Dendríticas , Microambiente Tumoral , Vacunas contra el Cáncer/inmunología , Microambiente Tumoral/inmunología , Microambiente Tumoral/efectos de los fármacos , Animales , Bacteriófago M13/inmunología , Bacteriófago M13/química , Ratones , Células Dendríticas/inmunología , Antígenos CD40/inmunología , Antígenos CD40/metabolismo , Ratones Endogámicos C57BL , Femenino , Línea Celular Tumoral , Factor Estimulante de Colonias de Granulocitos y Macrófagos , Antígenos de Neoplasias/inmunología , Humanos

RESUMEN

INTRODUCTION: There is a need for new therapies that can enhance response rates and broaden the number of cancer indications where immunotherapies provide clinical benefit. CD40 targeting therapies provide an opportunity to meet this need by promoting priming of tumor-specific T cells and reverting the suppressive tumor microenvironment. This is supported by emerging clinical evidence demonstrating the benefits of immunotherapy with CD40 antibodies in combination with standard of care chemotherapy. AREAS COVERED: This review is focused on the coming wave of next-generation CD40 agonists aiming to improve efficacy and safety, using new approaches and formats beyond monospecific antibodies. Further, the current understanding of the role of different CD40 expressing immune cell populations in the tumor microenvironment is reviewed. EXPERT OPINION: There are multiple promising next-generation approaches beyond monospecific antibodies targeting CD40 in immuno-oncology. Enhancing efficacy is the most important driver for this development, and approaches that maximize the ability of CD40 to both remodel the tumor microenvironment and boost the anti-tumor T cell response provide great opportunities to benefit cancer patients. Enhanced understanding of the role of different CD40 expressing immune cells in the tumor microenvironment may facilitate more efficient clinical development of these compounds.

Asunto(s)

Antígenos CD40 , Inmunoterapia , Neoplasias , Microambiente Tumoral , Humanos , Antígenos CD40/agonistas , Antígenos CD40/inmunología , Neoplasias/inmunología , Neoplasias/terapia , Neoplasias/tratamiento farmacológico , Animales , Microambiente Tumoral/inmunología

RESUMEN

B cells, despite their several unique functionalities, remain largely untapped for use as an adoptive cell therapy and are limited to in vitro use for antibody production. B cells can be easily sourced, they possess excellent lymphoid-homing capabilities, and they can act as antigen-presenting cells (APCs), offering an alternative to dendritic cells (DCs), which have shown limited efficacy in the clinical setting. Soluble factors such as IL-4 and anti-CD40 antibody can enhance the activation, survival, and antigen-presenting capabilities of B cells; however, it is difficult to attain sufficiently high concentrations of these biologics to stimulate B cells in vivo. Micropatches as Cell Engagers (MACE) are polymeric microparticles, surface functionalized with anti-CD40 and anti-IgM, which can attach to B cells and simultaneously engage multiple B-cell receptors (BCR) and CD40 receptors. Stimulation of these receptors through MACE, unlike free antibodies, enhanced the display of costimulatory molecules on the B-cell surface, increased B-cell viability, and improved antigen presentation by B cells to T cells in vitro. B-cell activation by MACE further synergized with soluble IL-4 and anti-CD40. MACE also elicited T-cell chemokine secretion by B cells. Upon intravenous adoptive transfer, MACE-bound B cells homed to the spleen and lymph nodes, key sites for antigen presentation to T cells. Adoptive transfer of MACE-B cells pulsed with the CD4+ and CD8+ epitopes of ovalbumin significantly delayed tumor progression in a murine subcutaneous EG7-OVA tumor model, demonstrating the functional benefit conferred to B cells by MACE.

Asunto(s)

Linfocitos B , Antígenos CD40 , Polímeros , Animales , Linfocitos B/inmunología , Ratones , Antígenos CD40/metabolismo , Antígenos CD40/inmunología , Polímeros/química , Receptores de Antígenos de Linfocitos B/metabolismo , Humanos , Linfocitos T/inmunología , Interleucina-4 , Ratones Endogámicos C57BL

RESUMEN

Anti-CD40 antibodies (Abs) have been shown to induce antitumor T-cell responses. We reported that the engineered agonistic anti-CD40 Ab (5C11, IgG4 isotype) recognized human CD40 antigen expressed on a human B lymphoblastoid cell line as well as on splenic cells isolated from humanized CD40 mice. Of note, a single high dosage of 5C11 was able to prohibit tumor growth in parallel with an increase in the population of infiltrated CD8+ T cells. Furthermore, the antitumor effects of 5C11 were enhanced in the presence of ß-glucan along with an increase in the population of infiltrated CD8+ T cells. In addition, the numbers of CD86+ TAMs and neutrophils were elevated in the combination of 5C11 and ß-glucan compared with either 5C11 or ß-glucan alone. Furthermore, the abundance of Faecalibaculum, one of the probiotics critical for tumor suppression, was obviously increased in the combination of 5C11 and ß-glucan-treated mice. These data reveal a novel mechanism of tumor suppression upon the combination treatment of 5C11 and ß-glucan and propose that the combination treatment of agonistic anti-human CD40 antibody 5C11 and ß-glucan could be a promising therapeutic strategy for cancer patients.

Asunto(s)

Antígenos CD40 , beta-Glucanos , Animales , Antígenos CD40/agonistas , Antígenos CD40/inmunología , Antígenos CD40/metabolismo , beta-Glucanos/farmacología , Ratones , Humanos , Anticuerpos Monoclonales/farmacología , Línea Celular Tumoral , Linfocitos T CD8-positivos/inmunología , Sinergismo Farmacológico

RESUMEN

An important challenge in the real-world management of patients with advanced clear-cell renal cell carcinoma (aRCC) is determining who might benefit from immune checkpoint blockade (ICB). Here we performed a comprehensive multiomics mapping of aRCC in the context of ICB treatment, involving discovery analyses in a real-world data cohort followed by validation in independent cohorts. We cross-connected bulk-tumor transcriptomes across >1,000 patients with validations at single-cell and spatial resolutions, revealing a patient-specific crosstalk between proinflammatory tumor-associated macrophages and (pre-)exhausted CD8+ T cells that was distinguished by a human leukocyte antigen repertoire with higher preference for tumoral neoantigens. A cross-omics machine learning pipeline helped derive a new tumor transcriptomic footprint of neoantigen-favoring human leukocyte antigen alleles. This machine learning signature correlated with positive outcome following ICB treatment in both real-world data and independent clinical cohorts. In experiments using the RENCA-tumor mouse model, CD40 agonism combined with PD1 blockade potentiated both proinflammatory tumor-associated macrophages and CD8+ T cells, thereby achieving maximal antitumor efficacy relative to other tested regimens. Thus, we present a new multiomics and spatial map of the immune-community architecture that drives ICB response in patients with aRCC.

Asunto(s)

Linfocitos T CD8-positivos , Carcinoma de Células Renales , Antígenos HLA , Inmunoterapia , Neoplasias Renales , Carcinoma de Células Renales/inmunología , Carcinoma de Células Renales/terapia , Carcinoma de Células Renales/genética , Carcinoma de Células Renales/patología , Humanos , Neoplasias Renales/inmunología , Neoplasias Renales/terapia , Neoplasias Renales/genética , Neoplasias Renales/patología , Animales , Inmunoterapia/métodos , Linfocitos T CD8-positivos/inmunología , Ratones , Antígenos HLA/inmunología , Antígenos HLA/genética , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inhibidores de Puntos de Control Inmunológico/farmacología , Aprendizaje Automático , Antígenos CD40/inmunología , Antígenos CD40/genética , Macrófagos Asociados a Tumores/inmunología , Transcriptoma , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Receptor de Muerte Celular Programada 1/inmunología , Femenino

RESUMEN

The parasite Leishmania resides as amastigotes within the macrophage parasitophorous vacuoles inflicting the disease Leishmaniasis. Leishmania selectively modulates mitogen-activated protein kinase (MAPK) phosphorylation subverting CD40-triggered anti-leishmanial functions of macrophages. The mechanism of any pathogen-derived molecule induced host MAPK modulation remains poorly understood. Herein, we show that of the fifteen MAPKs, LmjMAPK4 expression is higher in virulent L. major. LmjMAPK4- detected in parasitophorous vacuoles and cytoplasm- binds MEK-1/2, but not MKK-3/6. Lentivirally-overexpressed LmjMAPK4 augments CD40-activated MEK-1/2-ERK-1/2-MKP-1, but inhibits MKK3/6-p38MAPK-MKP-3, phosphorylation. A rationally-identified LmjMAPK4 inhibitor reinstates CD40-activated host-protective anti-leishmanial functions in L. major-infected susceptible BALB/c mice. These results identify LmjMAPK4 as a MAPK modulator at the host-pathogen interface and establish a pathogen-intercepted host receptor signaling as a scientific rationale for identifying drug targets.

Asunto(s)

Antígenos CD40 , Leishmania major , Leishmaniasis Cutánea , Macrófagos , Ratones Endogámicos BALB C , Transducción de Señal , Animales , Leishmania major/inmunología , Leishmania major/fisiología , Antígenos CD40/metabolismo , Ratones , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/parasitología , Macrófagos/inmunología , Macrófagos/parasitología , Humanos , Femenino , Fosforilación , Interacciones Huésped-Parásitos/inmunología , Sistema de Señalización de MAP Quinasas/inmunología

RESUMEN

The interaction between CD40 and CD40 ligand (CD40L) a crucial co-stimulatory signal for activating adaptive immune cells, has a noteworthy role in atherosclerosis. It is well-known that atherosclerosis is linked to immune inflammation in blood vessels. In atherosclerotic lesions, there is a multitude of proinflammatory cytokines, adhesion molecules, and collagen, as well as smooth muscle cells, macrophages, and T lymphocytes, particularly the binding of CD40 and CD40L. Therefore, research on inhibiting the CD40-CD40L system to prevent atherosclerosis has been ongoing for more than 30 years. However, it's essential to note that long-term direct suppression of CD40 or CD40L could potentially result in immunosuppression, emphasizing the critical role of the CD40-CD40L system in atherosclerosis. Thus, specifically targeting the CD40-CD40L interaction on particular cell types or their downstream signaling pathways may be a robust strategy for mitigating atherosclerosis, reducing potential side effects. This review aims to summarize the potential utility of the CD40-CD40L system as a viable therapeutic target for atherosclerosis.

Asunto(s)

Aterosclerosis , Ligando de CD40 , Humanos , Aterosclerosis/tratamiento farmacológico , Aterosclerosis/inmunología , Antígenos CD40/antagonistas & inhibidores , Antígenos CD40/metabolismo , Ligando de CD40/antagonistas & inhibidores , Ligando de CD40/metabolismo , Citocinas/metabolismo , Interleucina-2/metabolismo , Macrófagos/metabolismo

RESUMEN

CD40 is a member of the tumor necrosis factor receptor superfamily, and it is widely expressed on immune and non-immune cell types. The interaction between CD40 and the CD40 ligand (CD40L) plays an essential function in signaling, and the CD40/CD40L complex works as an immune checkpoint molecule. CD40 has become a therapeutic target, and a variety of agonistic/antagonistic anti-CD40 monoclonal antibodies (mAbs) have been developed. To better understand the mode of action of anti-CD40 mAbs, we determined the X-ray crystal structures of dacetuzumab (agonist) and bleselumab (antagonist) in complex with the extracellular domain of human CD40, respectively. The structure reveals that dacetuzumab binds to CD40 on the top of cysteine-rich domain 1 (CRD1), which is the domain most distant from the cell surface, and it does not compete with CD40L binding. The binding interface of bleselumab spread between CRD2 and CRD1, overlapping with the binding surface of the ligand. Our results offer important insights for future structural and functional studies of CD40 and provide clues to understanding the mechanism of biological response. These data can be applied to developing new strategies for designing antibodies with more therapeutic efficacy.

Asunto(s)

Anticuerpos Monoclonales Humanizados , Antígenos CD40 , Humanos , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales Humanizados/química , Anticuerpos Monoclonales Humanizados/inmunología , Sitios de Unión , Antígenos CD40/química , Antígenos CD40/inmunología , Antígenos CD40/metabolismo , Ligando de CD40/química , Ligando de CD40/metabolismo , Ligando de CD40/inmunología , Cristalografía por Rayos X , Modelos Moleculares , Unión Proteica , Conformación Proteica

RESUMEN

Xenotransplantation represents a possible solution to the organ shortage crisis and is an imminent clinical reality with long-term xenograft survival in pig-to-nonhuman primate (NHP) heart and kidney large animal models, and short-term success in recent human decedent and clinical studies. However, concerns remain about safe clinical translation of these results, given the inconsistency in published survival as well as key differences between preclinical procurement and immunosuppression and clinical standards-of-care. Notably, no studies of solid organ pig-to-NHP transplantation have achieved xenograft survival longer than one month without CD40/CD154 costimulatory blockade, which is not currently an FDA-approved immunosuppression strategy. We now present consistent survival in consecutive cases of pig-to-NHP kidney xenotransplantation, including long-term survival after >3 hours of xenograft cold preservation time as well as long-term survival using FDA-approved immunosuppression. These data provide critical supporting evidence for the safety and feasibility of clinical kidney xenotransplantation. Moreover, long-term survival without CD40/CD154 costimulatory blockade may provide important insights for immunosuppression regimens to be considered for first-in-human clinical trials.

Asunto(s)

Supervivencia de Injerto , Riñón , Animales , Humanos , Porcinos , Trasplante Heterólogo/métodos , Xenoinjertos , Terapia de Inmunosupresión/métodos , Ligando de CD40 , Antígenos CD40 , Rechazo de Injerto