RÉSUMÉ
ß C-S lyases (ß-CSLs; EC 4.4.1.8) are enzymes catalyzing the dissociation of ß carbon-sulfur bonds of cysteine S-conjugates to produce odorant metabolites with a free thiol group. These enzymes are increasingly studied for their role in flavor generation in a variety of food products, whether these processes occur directly in plants, by microbial ß-CSLs during fermentation, or in the mouth under the action of the oral microbiota. Microbial ß-CSLs react with sulfur aroma precursors present in beverages, vegetables, fruits, or aromatic herbs like hop but also potentially with some precursors formed through Maillard reactions in cooked foods such as meat or coffee. ß-CSLs from microorganisms like yeasts and lactic acid bacteria have been studied for their role in the release of polyfunctional thiols in wine and beer during fermentation. In addition, ß-CSLs from microorganisms of the human oral cavity were shown to metabolize similar precursors and to produce aroma in the mouth with an impact on retro-olfaction. This review summarizes the current knowledge on ß-CSLs involved in flavor generation with a focus on enzymes from microbial species present either in the fermentative processes or in the oral cavity. This paper highlights the importance of this enzyme family in the food continuum, from production to consumption, and offers new perspectives concerning the utilization of ß-CSLs as a flavor enhancer.
Sujet(s)
Fermentation , Aromatisants , Humains , Aromatisants/métabolisme , Carbon-sulfur lyases/métabolisme , Bactéries/enzymologie , Bactéries/métabolisme , GoûtRÉSUMÉ
Quantum computers are expected to outperform classical computers for specific problems in quantum chemistry. Such calculations remain expensive, but costs can be lowered through the partition of the molecular system. In the present study, partition was achieved with range-separated density functional theory (RS-DFT). The use of RS-DFT reduces both the basis set size and the active space size dependence of the ground state energy in comparison with the use of wave function theory (WFT) alone. The utilization of pair natural orbitals (PNOs) in place of canonical molecular orbitals (MOs) results in more compact qubit Hamiltonians. To test this strategy, a basis-set independent framework, known as multiresolution analysis (MRA), was employed to generate PNOs. Tests were conducted with the variational quantum eigensolver for a number of molecules. The results show that the proposed approach reduces the number of qubits needed to reach a target energy accuracy.
RÉSUMÉ
BACKGROUND: Allogeneic hematopoietic stem cell transplantation (HSCT) remains the standard of care for chemotherapy-refractory leukemia patients, but cure rates are still dismal. To prevent leukemia relapse following HSCT, we aim to improve the early graft-versus-leukemia effect mediated by natural killer (NK) cells. Our approach is based on the adoptive transfer of Therapeutic Inducers of Natural Killer cell Killing (ThINKK). ThINKK are expanded and differentiated from HSC, and exhibit blood plasmacytoid dendritic cell (pDC) features. We previously demonstrated that ThINKK stimulate NK cells and control acute lymphoblastic leukemia (ALL) development in a preclinical mouse model of HSCT for ALL. Here, we assessed the cellular identity of ThINKK and investigated their potential to activate allogeneic T cells. We finally evaluated the effect of immunosuppressive drugs on ThINKK-NK cell interaction. METHODS: ThINKK cellular identity was explored using single-cell RNA sequencing and flow cytometry. Their T-cell activating potential was investigated by coculture of allogeneic T cells and antigen-presenting cells in the presence or the absence of ThINKK. A preclinical human-to-mouse xenograft model was used to evaluate the impact of ThINKK injections on graft-versus-host disease (GvHD). Finally, the effect of immunosuppressive drugs on ThINKK-induced NK cell cytotoxicity against ALL cells was tested. RESULTS: The large majority of ThINKK shared the key characteristics of canonical blood pDC, including potent type-I interferon (IFN) production following Toll-like receptor stimulation. A minor subset expressed some, although not all, markers of other dendritic cell populations. Importantly, while ThINKK were not killed by allogeneic T or NK cells, they did not increase T cell proliferation induced by antigen-presenting cells nor worsened GvHD in vivo. Finally, tacrolimus, sirolimus or mycophenolate did not decrease ThINKK-induced NK cell activation and cytotoxicity. CONCLUSION: Our results indicate that ThINKK are type I IFN producing cells with low T cell activation capacity. Therefore, ThINKK adoptive immunotherapy is not expected to increase the risk of GvHD after allogeneic HSCT. Furthermore, our data predict that the use of tacrolimus, sirolimus or mycophenolate as anti-GvHD prophylaxis regimen will not decrease ThINKK therapeutic efficacy. Collectively, these preclinical data support the testing of ThINKK immunotherapy in a phase I clinical trial.
Sujet(s)
Transplantation de cellules souches hématopoïétiques , Cellules tueuses naturelles , Cellules tueuses naturelles/immunologie , Cellules tueuses naturelles/métabolisme , Cellules tueuses naturelles/effets des médicaments et des substances chimiques , Humains , Transplantation de cellules souches hématopoïétiques/méthodes , Animaux , Souris , Transplantation homologue , Cellules dendritiques/immunologie , Cellules dendritiques/métabolisme , Femelle , Maladie du greffon contre l'hôte/prévention et contrôleRÉSUMÉ
ABSTRACT: Acute lymphoblastic leukemia (ALL) arises from the uncontrolled proliferation of B-cell precursors (BCP-ALL) or T cells (T-ALL). Current treatment protocols obtain high cure rates in children but are based on toxic polychemotherapy. Novel therapies are urgently needed, especially in relapsed/refractory (R/R) disease, high-risk (HR) leukemias and T-ALL, in which immunotherapy approaches remain scarce. Although the interleukin-7 receptor (IL-7R) plays a pivotal role in ALL development, no IL-7R-targeting immunotherapy has yet reached clinical application in ALL. The IL-7Rα chain (CD127)-targeting IgG4 antibody lusvertikimab (LUSV; formerly OSE-127) is a full antagonist of the IL-7R pathway, showing a good safety profile in healthy volunteers. Here, we show that â¼85% of ALL cases express surface CD127. We demonstrate significant in vivo efficacy of LUSV immunotherapy in a heterogeneous cohort of BCP- and T-ALL patient-derived xenografts (PDX) in minimal residual disease (MRD) and overt leukemia models, including R/R and HR leukemias. Importantly, LUSV was particularly effective when combined with polychemotherapy in a phase 2-like PDX study with CD127high samples leading to MRD-negativity in >50% of mice treated with combination therapy. Mechanistically, LUSV targeted ALL cells via a dual mode of action comprising direct IL-7R antagonistic activity and induction of macrophage-mediated antibody-dependent cellular phagocytosis (ADCP). LUSV-mediated in vitro ADCP levels significantly correlated with CD127 expression levels and the reduction of leukemia burden upon treatment of PDX animals in vivo. Altogether, through its dual mode of action and good safety profile, LUSV may represent a novel immunotherapy option for any CD127+ ALL, particularly in combination with standard-of-care polychemotherapy.
Sujet(s)
Tests d'activité antitumorale sur modèle de xénogreffe , Animaux , Humains , Souris , Récepteurs à l'interleukine-7/antagonistes et inhibiteurs , Souris SCID , Phagocytose/effets des médicaments et des substances chimiques , Sous-unité alpha du récepteur à l'interleukine-7 , Leucémie-lymphome lymphoblastique à précurseurs B et T/traitement médicamenteux , Leucémie-lymphome lymphoblastique à précurseurs B et T/anatomopathologie , Leucémie-lymphome lymphoblastique à précurseurs B et T/immunologie , Femelle , Souris de lignée NOD , Anticorps monoclonaux humanisés/usage thérapeutique , Anticorps monoclonaux humanisés/pharmacologie , Lignée cellulaire tumorale , Antinéoplasiques immunologiques/pharmacologie , Antinéoplasiques immunologiques/usage thérapeutiqueRÉSUMÉ
Glutathione transferases are xenobiotic-metabolizing enzymes with both glutathione-conjugation and ligandin roles. GSTs are present in chemosensory tissues and fluids of the nasal/oral cavities where they protect tissues from exogenous compounds, including food molecules. In the present study, we explored the presence of the omega-class glutathione transferase (GSTO1) in the rat oral cavity. Using immunohistochemistry, GSTO1 expression was found in taste bud cells of the tongue epithelium and buccal cells of the oral epithelium. Buccal and lingual extracts exhibited thiol-transferase activity (4.9 ± 0.1 and 1.8 ± 0.1 µM/s/mg, respectively). A slight reduction from 4.9 ± 0.1 to 4.2 ± 0.1 µM/s/mg (p < 0.05; Student's t test) was observed in the buccal extract with 100 µM GSTO1-IN-1, a specific inhibitor of GSTO1. RnGSTO1 exhibited the usual activities of omega GSTs, i.e., thiol-transferase (catalytic efficiency of 8.9 × 104 M-1·s-1), and phenacyl-glutathione reductase (catalytic efficiency of 8.9 × 105 M-1·s-1) activities, similar to human GSTO1. RnGSTO1 interacts with food phytochemicals, including bitter compounds such as luteolin (Ki = 3.3 ± 1.9 µM). Crystal structure analysis suggests that luteolin most probably binds to RnGSTO1 ligandin site. Our results suggest that GSTO1 could interact with food phytochemicals in the oral cavity.
Sujet(s)
Glutathione transferase , Lutéoline , Rats , Animaux , Humains , Glutathione transferase/métabolisme , Muqueuse de la bouche/métabolisme , Thiols , Glutathion/métabolismeRÉSUMÉ
The inflammatory response is a key mechanism for the elimination of injurious agents but must be tightly controlled to prevent additional tissue damage and progression to persistent inflammation. C-type lectin receptors expressed mostly by myeloid cells play a crucial role in the regulation of inflammation by recognizing molecular patterns released by injured tissues. We recently showed that the C-type lectin receptor CLEC-1 is able to recognize necrotic cells. However, its role in the acute inflammatory response following tissue damage had not yet been investigated. We show in this study, in a mouse model of liver injury induced by acetaminophen intoxication, that Clec1a deficiency enhances the acute immune response with increased expression of Il1b, Tnfa, and Cxcl2 and higher infiltration of activated neutrophils into the injured organ. Furthermore, we demonstrate that Clec1a deficiency exacerbates tissue damage via CXCL2-dependent neutrophil infiltration. In contrast, we observed that the lack of CLEC-1 limits CCL2 expression and the accumulation, beyond the peak of injury, of monocyte-derived macrophages. Mechanistically, we found that Clec1a-deficient dendritic cells increase the expression of Il1b, Tnfa, and Cxcl2 in response to necrotic cells, but decrease the expression of Ccl2. Interestingly, treatment with an anti-human CLEC-1 antagonist mAb recapitulates the exacerbation of acute immunopathology observed by genetic loss of Clec1a in a preclinical humanized mouse model. To conclude, our results demonstrate that CLEC-1 is a death receptor limiting the acute inflammatory response following injury and represents a therapeutic target to modulate immunity.
Sujet(s)
Inflammation , Granulocytes neutrophiles , Souris , Animaux , Cellules myéloïdes , Macrophages , Foie/métabolisme , Lectines de type C/métabolismeRÉSUMÉ
OBJECTIVE: While the involvement of IL-7/IL-7R axis in pSS has been described in relation to T cells, little is known about the contribution of this pathway in relationship with other immune cells, and its implication in autoimmunity. Using high-content multiomics data, we aimed at characterizing IL-7R expressing cells and the involvement of IL-7/IL-7R pathway in pSS pathophysiology. METHODS: An IL-7 signature established using RNA-sequencing of human PBMCs incubated with IL-7 was applied to 304 pSS patients, and on RNA-Seq datasets from tissue biopsies. High-content immunophenotyping using flow and imaging mass cytometry was developed to characterize peripheral and in situ IL-7R expression. RESULTS: We identified a blood 4-gene IL-7 module (IKZF4, KIAA0040, PGAP1 and SOS1) associated with anti-SSA/Ro positiveness in patients as well as disease activity, and a tissue 5-gene IL-7 module (IL7R, PCED1B, TNFSF8, ADAM19, MYBL1) associated with infiltration severity. We confirmed expression of IL-7R on T cells subsets, and further observed upregulation of IL-7R on double-negative (DN) B cells, and especially DN2 B cells. IL-7R expression was increased in pSS compared to sicca patients with variations seen according to the degree of infiltration. When expressed, IL-7R was mainly found on epithelial cells, CD4+ and CD8+ T cells, switched memory B cells, DN B cells and M1 macrophages. CONCLUSION: This exhaustive characterization of the IL-7/IL-7R pathway in pSS pathophysiology established that two IL-7 gene modules discriminate pSS patients with a high IL-7 axis involvement. Their use could guide the implementation of an anti-IL-7R targeted therapy in a precision medicine approach.
RÉSUMÉ
Introduction: Tumor Associated Macrophages (TAM) are a major component of the tumor environment and their accumulation often correlates with poor prognosis by contributing to local inflammation, inhibition of anti-tumor immune response and resistance to anticancer treatments. In this study, we thus investigated the anti-cancer therapeutic interest to target ChemR23, a receptor of the resolution of inflammation expressed by macrophages, using an agonist monoclonal antibody, αChemR23. Methods: Human GM-CSF, M-CSF and Tumor Associated Macrophage (TAM)-like macrophages were obtained by incubation of monocytes from healthy donors with GM-CSF, M-CSF or tumor cell supernatants (Breast cancer (BC) or malignant pleural mesothelioma (MPM) cells). The effects of αChemR23 on macrophages were studied at the transcriptomic, protein and functional level. Datasets from The Cancer Genome Atlas (TCGA) were used to study CMKLR1 expression, coding for ChemR23, in BC and MPM tumors. In vivo, αChemR23 was evaluated on overall survival, metastasis development and transcriptomic modification of the metastatic niche using a model of resected triple negative breast cancer. Results: We show that ChemR23 is expressed at higher levels in M-CSF and tumor cell supernatant differentiated macrophages (TAM-like) than in GM-CSF-differentiated macrophages. ChemR23 activation triggered by αChemR23 deeply modulates M-CSF and TAM-like macrophages including profile of cell surface markers, cytokine secretion, gene mRNA expression and immune functions. The expression of ChemR23 coding gene (CMKLR1) strongly correlates to TAM markers in human BC tumors and MPM and its histological detection in these tumors mainly corresponds to TAM expression. In vivo, treatment with αChemR23 agonist increased mouse survival and decreased metastasis occurrence in a model of triple-negative BC in correlation with modulation of TAM phenotype in the metastatic niche. Conclusion: These results open an attractive opportunity to target TAM and the resolution of inflammation pathways through ChemR23 to circumvent TAM pro-tumoral effects.
Sujet(s)
Tumeurs du sein , Carcinomes , Récepteurs aux chimiokines , Animaux , Femelle , Humains , Souris , Carcinomes/métabolisme , Facteur de stimulation des colonies de granulocytes et de macrophages/métabolisme , Inflammation/métabolisme , Facteur de stimulation des colonies de macrophages/métabolisme , Macrophages , Phénotype , Récepteurs aux chimiokines/métabolismeRÉSUMÉ
The annual "Antibody Industrial Symposium", co-organized by LabEx MAbImprove and MabDesign, held its 10th anniversary edition in Montpellier, France, on June 28-29, 2022. The meeting focused on new results and concepts in antibody engineering (naked, mono- or multi-specific, conjugated to drugs or radioelements) and also on new cell-based therapies, such as chimeric antigenic receptor (CAR)-T cells. The symposium, which brought together scientists from academia and industry, also addressed issues concerning the production of these molecules and cells, and the necessary steps to ensure a strong intellectual property protection of these new molecules and approaches. These two days of exchanges allowed a rich discussion among the various actors in the field of therapeutic antibodies.
Sujet(s)
Anticorps monoclonaux , Immunothérapie adoptive , Anticorps monoclonaux/usage thérapeutique , FranceRÉSUMÉ
OSE-127 is a humanized mAb targeting the IL-7Rα-chain (CD127), under development for inflammatory and autoimmune disease treatment. It is a strict antagonist of the IL-7R pathway, is not internalized by target cells, and is noncytotoxic. In this work, a first-in-human, phase I, randomized, double-blind, placebo-controlled, single-center study was carried out to determine the safety, pharmacokinetics, pharmacodynamics, and immunogenicity of OSE-127 administration. Sixty-three healthy subjects were randomly assigned to nine groups: six single ascending dose groups with i.v. administration (0.002-10 mg/kg), a single s.c. treatment group (1 mg/kg), and two double i.v. injection groups (6 or 10 mg/kg). Subjects were followed during <146 d. OSE-127's pharmacokinetic half-life after a single dose increased from 4.6 (1 mg/kg) to 11.7 d (10 mg/kg) and, after a second dose, from 12.5 (6 mg/kg) to 16.25 d (10 mg/kg). Receptor occupancy was ≥95% at doses ≥0.02 mg/kg, and this saturation level was maintained >100 d after two i.v. infusions at 10 mg/kg. IL-7 consumption was inhibited by OSE-127 administration, as demonstrated by a decreased IL-7 pathway gene signature in peripheral blood cells and by ex vivo T lymphocyte restimulation experiments. OSE-127 was well tolerated, with no evidence of cytokine-release syndrome and no significant alteration of blood lymphocyte counts or subset populations. Altogether, the observed lack of significant lymphopenia or serious adverse events, concomitant with the dose-dependent inhibition of IL-7 consumption by target cells, highlights that OSE-127 may show clinical activity in IL-7R pathway-involved diseases.
Sujet(s)
Anticorps monoclonaux , Interleukine-7 , Humains , Anticorps monoclonaux/usage thérapeutique , Volontaires sains , Anticorps monoclonaux humanisés/pharmacologie , Méthode en double aveugle , Relation dose-effet des médicamentsRÉSUMÉ
Tumors exploit numerous immune checkpoints, including those deployed by myeloid cells to curtail antitumor immunity. Here, we show that the C-type lectin receptor CLEC-1 expressed by myeloid cells senses dead cells killed by programmed necrosis. Moreover, we identified Tripartite Motif Containing 21 (TRIM21) as an endogenous ligand overexpressed in various cancers. We observed that the combination of CLEC-1 blockade with chemotherapy prolonged mouse survival in tumor models. Loss of CLEC-1 reduced the accumulation of immunosuppressive myeloid cells in tumors and invigorated the activation state of dendritic cells (DCs), thereby increasing T cell responses. Mechanistically, we found that the absence of CLEC-1 increased the cross-presentation of dead cell-associated antigens by conventional type-1 DCs. We identified antihuman CLEC-1 antagonist antibodies able to enhance antitumor immunity in CLEC-1 humanized mice. Together, our results demonstrate that CLEC-1 acts as an immune checkpoint in myeloid cells and support CLEC-1 as a novel target for cancer immunotherapy.
Sujet(s)
Cross-priming , Tumeurs , Souris , Animaux , Présentation d'antigène , Immunothérapie , Cellules dendritiques , Tumeurs/thérapieRÉSUMÉ
The BAG3- and SIRPα- mediated pathways trigger distinct cellular targets and signaling mechanisms in pancreatic cancer microenvironment. To explore their functional connection, we investigated the effects of their combined blockade on cancer growth in orthotopic allografts of pancreatic cancer mt4-2D cells in immunocompetent mice. The anti-BAG3 + anti-SIRPα mAbs treatment inhibited (p = 0.007) tumor growth by about the 70%; also the number of metastatic lesions was decreased, mostly by the effect of the anti-BAG3 mAb. Fibrosis and the expression of the CAF activation marker α-SMA were reduced by about the 30% in animals treated with anti-BAG3 mAb compared to untreated animals, and appeared unaffected by treatment with the anti-SIRPα mAb alone; however, the addition of anti-SIRPα to anti-BAG3 mAb in the combined treatment resulted in a > 60% (p < 0.0001) reduction of the fibrotic area and a 70% (p < 0.0001) inhibition of CAF α-SMA positivity. Dendritic cells (DCs) and CD8+ lymphocytes, hardly detectable in the tumors of untreated animals, were modestly increased by single treatments, while were much more clearly observable (p < 0.0001) in the tumors of the animals subjected to the combined treatment. The effects of BAG3 and SIRPα blockade do not simply reflect the sum of the effects of the single blockades, indicating that the two pathways are connected by regulatory interactions and suggesting, as a proof of principle, the potential therapeutic efficacy of a combined BAG3 and SIRPα blockade in pancreatic cancer.
RÉSUMÉ
A numerous number of positive and negative signals via various molecules modulate T-cell activation. Within the various transmembrane proteins, SIRPγ is of interest since it is not expressed in rodents. SIRPγ interaction with CD47 is reevaluated in this study. Indeed, we show that the anti-SIRPγ mAb clone LSB2.20 previously used by others has not been appropriately characterized. We reveal that the anti-SIRPα clone KWAR23 is a Pan anti-SIRP mAb which efficiently blocks SIRPα and SIRPγ interactions with CD47. We show that SIRPγ expression on T cells varies with their differentiation and while being expressed on Tregs, is not implicated in their suppressive functions. SIRPγ spatial reorganization at the immune synapse is independent of its interaction with CD47. In vitro SIRPα-γ/CD47 blockade with KWAR23 impairs IFN-γ secretion by chronically activated T cells. In vivo in a xeno-GvHD model in NSG mice, the SIRPγ/CD47 blockade with the KWAR23 significantly delays the onset of the xeno-GvHD and deeply impairs human chimerism. In conclusion, we have shown that T-cell interaction with CD47 is of importance notably in chronic stimulation.
Sujet(s)
Antigènes de différenciation/métabolisme , Antigènes CD47/métabolisme , Maladie du greffon contre l'hôte/immunologie , Activation des lymphocytes/effets des médicaments et des substances chimiques , Muromonab-CD3/administration et posologie , Récepteurs immunologiques/métabolisme , Transduction du signal/effets des médicaments et des substances chimiques , Lymphocytes T/immunologie , Animaux , Antigènes de différenciation/génétique , Antigènes de différenciation/immunologie , Donneurs de sang , Antigènes CD47/génétique , Modèles animaux de maladie humaine , Femelle , Techniques de knock-in de gènes , Techniques de knock-out de gènes , Volontaires sains , Hétérogreffes , Humains , Cellules Jurkat , Activation des lymphocytes/génétique , Mâle , Souris , Muromonab-CD3/immunologie , Récepteurs immunologiques/génétique , Récepteurs immunologiques/immunologie , Transduction du signal/génétiqueRÉSUMÉ
Inflammation is a fundamental physiological response orchestrated by innate immune cells to restore tissue homeostasis. Specialized pro-resolving mediators (SPMs) are involved in active resolution of inflammation but when inflammation is incomplete, chronic inflammation creates a favorable environment that fuels carcinogenesis and cancer progression. Conventional cancer therapy also strengthens cancer-related inflammation by inducing massive tumor cell death that activate surrounding immune-infiltrating cells such as tumor-associated macrophages (TAMs). Macrophages are key actors of both inflammation and its active resolution due to their plastic phenotype. In line with this high plasticity, macrophages can be hijacked by cancer cells to support tumor progression and immune escape, or therapy resistance. Impaired resolution of cancer-associated inflammation supported by TAMs may thus reinforces tumor progression. From this perspective, recent evidence suggests that stimulating macrophage's pro-resolving functions using SPMs can promote inflammation resolution in cancer and improve anticancer treatments. Thus, TAMs' re-education toward an antitumor phenotype by using SPMs opens a new line of attack in cancer treatment. Here, we review SPMs' anticancer capacities with special attention regarding their effects on TAMs. We further discuss how this new therapeutic approach could be envisioned in cancer therapy.
Sujet(s)
Médiateurs de l'inflammation/immunologie , Inflammation/immunologie , Tumeurs/immunologie , Macrophages associés aux tumeurs/immunologie , Animaux , HumainsRÉSUMÉ
Brazzein is a small sweet-tasting protein found in the red berries of a West African evergreen shrub, Pentadiplandra brazzeana Baillon. Brazzein is highly soluble and stable over a large pH range and at high temperatures, which are characteristics that suggest its use as a natural sweetener. However, Pentadiplandra brazzeana culture is difficult at a large scale, limiting the natural source of brazzein. Heterologous expression of brazzein has been established in numerous systems, including bacteria, yeast, and transgenic plants. Brazzein requires four disulfide bonds to be active in eliciting an intense sweet taste, and the yeast Pichia pastoris appears to be one of the best options for obtaining functional brazzein in high quantities. Employing yeast secretion in the culture medium allows us to obtain fully active brazzein and facilitate purification later. To increase yeast secretion, we compared seven different signal peptides to successfully achieve brazzein secretion using the yeast P. pastoris. The brazzein proteins corresponding to these signal peptides elicited activation of the sweet taste receptor functionally expressed in a cellular assay. Among these tested signal peptides, three resulted in the secretion of brazzein at high levels.
RÉSUMÉ
OBJECTIVE: Primary Sjögren's syndrome (SS) is characterized by a lymphocytic infiltration of salivary glands (SGs) and the presence of an interferon (IFN) signature. SG epithelial cells (SGECs) play an active role in primary SS pathophysiology. We undertook this study to examine the interactions between SGECs and T cells in primary SS and the role of the interleukin-7 (IL-7)/IFN axis. METHODS: Primary cultured SGECs from control subjects and patients with primary SS were stimulated with poly(I-C), IFNα, or IFNγ. T cells were sorted from blood and stimulated with IL-7. CD25 expression was assessed by flow cytometry. SG explants were cultured for 4 days with anti-IL-7 receptor (IL-7R) antagonist antibody (OSE-127), and transcriptomic analysis was performed using the NanoString platform. RESULTS: Serum IL-7 level was increased in patients with primary SS compared to controls and was associated with B cell biomarkers. IL7R expression was decreased in T cells from patients with primary SS compared to controls. SGECs stimulated with poly(I-C), IFNα, or IFNγ secreted IL-7. IL-7 stimulation increased the activation of T cells, as well as IFNγ secretion. Transcriptomic analysis of SG explants showed a correlation between IL7 and IFN expression. Finally, explants cultured with anti-IL-7R antibody showed decreased IFN-stimulated gene expression. CONCLUSION: These results suggest the presence of an IL-7/IFNγ amplification loop involving SGECs and T cells in primary SS. IL-7 was secreted by SGECs stimulated with type I or type II IFN and, in turn, activated T cells that secrete type II IFN. An anti-IL-7R antibody decreased the IFN signature in T cells in primary SS and could be of therapeutic interest.
Sujet(s)
Cellules épithéliales/métabolisme , Interféron alpha/pharmacologie , Interféron gamma/pharmacologie , Interleukine-7/pharmacologie , Glandes salivaires/métabolisme , Syndrome de Gougerot-Sjögren/métabolisme , Lymphocytes T/métabolisme , Adulte , Sujet âgé , Cellules cultivées , Cellules épithéliales/effets des médicaments et des substances chimiques , Femelle , Humains , Sous-unité alpha du récepteur à l'interleukine-7/immunologie , Mâle , Adulte d'âge moyen , Glandes salivaires/effets des médicaments et des substances chimiques , Transduction du signal/effets des médicaments et des substances chimiques , Lymphocytes T/effets des médicaments et des substances chimiquesRÉSUMÉ
T cell exclusion causes resistance to cancer immunotherapies via immune checkpoint blockade (ICB). Myeloid cells contribute to resistance by expressing signal regulatory protein-α (SIRPα), an inhibitory membrane receptor that interacts with ubiquitous receptor CD47 to control macrophage phagocytosis in the tumor microenvironment. Although CD47/SIRPα-targeting drugs have been assessed in preclinical models, the therapeutic benefit of selectively blocking SIRPα, and not SIRPγ/CD47, in humans remains unknown. We report a potent synergy between selective SIRPα blockade and ICB in increasing memory T cell responses and reverting exclusion in syngeneic and orthotopic tumor models. Selective SIRPα blockade stimulated tumor nest T cell recruitment by restoring murine and human macrophage chemokine secretion and increased anti-tumor T cell responses by promoting tumor-antigen crosspresentation by dendritic cells. However, nonselective SIRPα/SIRPγ blockade targeting CD47 impaired human T cell activation, proliferation, and endothelial transmigration. Selective SIRPα inhibition opens an attractive avenue to overcoming ICB resistance in patients with elevated myeloid cell infiltration in solid tumors.
Sujet(s)
Mémoire immunologique , Immunothérapie , Tumeurs expérimentales de la mamelle/thérapie , Protéines tumorales/immunologie , Récepteurs immunologiques/immunologie , Lymphocytes T/immunologie , Animaux , Femelle , Tumeurs expérimentales de la mamelle/immunologie , Tumeurs expérimentales de la mamelle/anatomopathologie , Souris , Souris de lignée BALB C , Protéines tumorales/génétique , Récepteurs immunologiques/génétique , Lymphocytes T/anatomopathologieRÉSUMÉ
Vertebrate odorant-binding proteins (OBPs) are small soluble proteins abundantly secreted in the olfactory mucus of many animal species, including humans. Vertebrate OBPs reversibly bind odorant molecules with micromolar range affinities. Although their physiological role is not clearly understood, OBPs are proposed to carry airborne odorants toward membrane olfactory receptors through the nasal mucus. Measurements of odorant-OBP interactions and structural studies require a large amount of pure OBPs devoid of ligands. The bacterial expression system is the first choice for expressing vertebrate OBPs used in our laboratory and others. This system generally produces OBPs in large amounts without major problems. In this chapter, we describe the milligram-scale production of recombinant pig OBP1 (pOBP1) in E. coli. The different steps of expression and purification are presented and discussed. Protocols for secondary structures investigation by circular dichroism and binding properties of the recombinant protein are also provided. More generally, these approaches can be used to produce and characterize any vertebrate OBPs for use in functional and structural studies.
Sujet(s)
Récepteurs olfactifs , Animaux , Protéines de transport , Escherichia coli/génétique , Escherichia coli/métabolisme , Protéines d'insecte , Odorisants , Récepteurs olfactifs/génétique , Récepteurs olfactifs/métabolisme , Suidae , Vertébrés/métabolismeRÉSUMÉ
IL-7 is an important cytokine for T cell lymphopoiesis. Blockade of the IL-7 signaling pathway has been shown to induce long-term graft survival or graft tolerance in murine transplant models through inhibiting T cell homeostasis and favoring immunoregulation. In this study, we assessed for the first time the effects of a blocking anti-human cluster of differentiation 127 (CD127) mAb administered in combination with low-dose tacrolimus or thymoglobulin in a life-sustaining kidney allograft model in baboons. Contrary to our expectation, the addition of an anti-CD127 mAb to the treatment protocols did not prolong graft survival compared to low-dose tacrolimus alone or thymoglobulin alone. Anti-CD127 mAb administration led to full CD127 receptor occupancy during the follow-up period. However, all treated animals lost their kidney graft between 1 week and 2 weeks after transplantation. Unlike in rodents, in nonhuman primates, anti-CD127 mAb treatment does not decrease the absolute numbers of lymphocyte and lymphocyte subsets and does not effectively inhibit postdepletional T cell proliferation and homeostasis, suggesting that IL-7 is not a limiting factor for T cell homeostasis in primates.
Sujet(s)
Anticorps monoclonaux/pharmacologie , Rejet du greffon/traitement médicamenteux , Survie du greffon/effets des médicaments et des substances chimiques , Sous-unité alpha du récepteur à l'interleukine-7/immunologie , Transplantation rénale/effets indésirables , Déplétion lymphocytaire/méthodes , Récepteurs à l'interleukine-7/antagonistes et inhibiteurs , Animaux , Rejet du greffon/étiologie , Rejet du greffon/anatomopathologie , Survie du greffon/immunologie , Papio , Complications postopératoiresRÉSUMÉ
Odorant-binding proteins (OBPs) are small soluble proteins that are thought to transport hydrophobic odorants across the aqueous sensillar lymph to olfactory receptors. A recent study revealed that OBP28a, one of the most abundant Drosophila OBPs, is not required for odorant transport, but acts in buffering rapid odour variation in the odorant environment. To further unravel and decipher its functional role, we expressed recombinant OBP28a and characterized its binding specificity. Using a fluorescent binding assay, we found that OBP28a binds a restricted number of floral-like chemicals, including ß-ionone, with an affinity in the micromolar range. We solved the X-ray crystal structure of OBP28a, which showed extensive conformation changes upon ligand binding. Mutant flies genetically deleted for the OBP28a gene showed altered responses to ß-ionone at a given concentration range, supporting its essential role in the detection of specific compounds present in the natural environment of the fly.