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1.
Theranostics ; 14(16): 6249-6267, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-39431011

RESUMEN

Rationale: CD39, a key ectonucleotidase that drives adenosine production, acts as a critical immunosuppressive checkpoint in cancer. Although it has shown promise as a therapeutic target, clinical trials are demonstrating the need for more potent targeting approaches. This need is driving innovation towards the development of novel antibodies and the exploration of strategic combinations with a range of immunotherapies. Methods: An anti-CD39 nanobody was screened and tested for its affinity and binding ability using biolayer interferometry, ELISA and flow cytometry. Blocking ability against soluble and membrane-bound CD39 was measured after CD39 blockade. Internalization was detected using immunofluorescence. The reversal of T-cell function by the anti-CD39 antibody was assessed by CFSE-based T-cell proliferation, CD25 expression and IFN-γ secretion. The in vivo function of tumor growth inhibition was further tested in a mouse model and we also tested the phenotype of immune cells after CD39 antibody administration from tumor tissue, draining lymph nodes and peripheral blood. We inserted the antibody sequence into the chimeric antigen receptor (CAR) construct to induce MSLN CAR-T cells to secret the CD39 antibody, and the efficacy was measured in xenograft models of ovarian cancer. Results: We screened human CD39 antibodies using a VHH library and developed a single-epitope anti-CD39 nanobody, named huCD39 mAb, with high affinity and potent binding and blocking ability. The huCD39 mAb was internalized in a time-dependent manner. The in vitro study revealed that the huCD39 mAb was highly effective in enhancing T-cell proliferation and functionality. In vivo, the huCD39 mAb showed significant anti-tumor efficacy in an immunocompetent mouse model. Flow cytometry analysis demonstrated downregulated CD39 expression in immune cells after antibody administration. We also observed increased CD39 expression in ovarian cancer tissue and in activated CAR T cells. Subsequently, we developed a type of MSLN CAR-T cells secreting huCD39 mAb which showed effective eradication or inhibition in ovarian tumor xenografts. Conclusions: A novel huCD39 mAb with strong blocking ability against human CD39 and potent inhibition of tumor growth has been developed. Furthermore, a modified huCD39 mAb-secreting CAR-T cell has been generated, exhibiting superior efficacy against ovarian cancer. This provides a promising strategy for optimizing immunotherapies in ovarian cancer and potentially other malignancies.


Asunto(s)
Apirasa , Neoplasias Ováricas , Receptores Quiméricos de Antígenos , Anticuerpos de Dominio Único , Linfocitos T , Animales , Neoplasias Ováricas/inmunología , Neoplasias Ováricas/terapia , Femenino , Apirasa/inmunología , Apirasa/metabolismo , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/metabolismo , Anticuerpos de Dominio Único/inmunología , Ratones , Humanos , Línea Celular Tumoral , Linfocitos T/inmunología , Linfocitos T/metabolismo , Inmunoterapia Adoptiva/métodos , Mesotelina , Ensayos Antitumor por Modelo de Xenoinjerto , Antígenos CD/inmunología , Antígenos CD/metabolismo , Proliferación Celular
2.
Int Immunopharmacol ; 140: 112743, 2024 Oct 25.
Artículo en Inglés | MEDLINE | ID: mdl-39094356

RESUMEN

BACKGROUND: Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease characterized by immune mechanisms dysregulation, leading to the production of diverse autoantibodies. However, the immune pathways underlying B-cell function and phenotypic abnormalities related to SLE pathogenesis remain incompletely understood. OBJECTIVE: To explore new markers of SLE activity and potential targets for SLE immunotherapy. METHODS: Collect peripheral blood mononuclear cells (PBMCs) from SLE patients and healthy controls (HC). Use flow cytometry to detect CD39 and CD73 expression on B cell subsets and enzyme-linked immunosorbent assay (ELISA) to measure adenosine (ADO) concentrations in SLE patients' serum. Compare CD39+CD73+ B cell subsets frequency and ADO concentrations in SLE patients and HC group. Additionally, analyze the correlation between CD39+CD73+ B cell subsets frequency and clinical laboratory parameters. RESULTS: CD39 and CD73 are simultaneously highly expressed on CD19+ B cell subsets, with significantly lower frequency of CD39+CD73+ B cell subsets in SLE patients compared to HC group. This frequency negatively correlates with Systemic Lupus Erythematosus Disease Activity Index (SLEDAI), C-reactive protein (CRP), and anti-double-stranded DNA (anti-dsDNA) antibodies, while positively correlating with IgM and prothrombin time (PT). Additionally, the frequency of CD39+CD73+ B cell subsets is significantly negatively correlated with IL-6 and IFN-α. In vitro cell experiments demonstrate that adenosine significantly inhibits R848-induced inflammatory cytokine production in a dose-dependent manner. CONCLUSION: The frequency of CD39+CD73+ B cell subsets of SLE patients is decreased, correlating with clinical laboratory parameters and disease activity. Simultaneously, ADO concentration in the patients' serum is reduced. The CD39+CD73+ B cell/ADO pathway may represent a novel immunotherapy strategy for SLE.


Asunto(s)
5'-Nucleotidasa , Adenosina , Apirasa , Lupus Eritematoso Sistémico , Humanos , Lupus Eritematoso Sistémico/inmunología , Lupus Eritematoso Sistémico/sangre , Apirasa/metabolismo , Apirasa/inmunología , 5'-Nucleotidasa/inmunología , 5'-Nucleotidasa/metabolismo , Femenino , Masculino , Adulto , Adenosina/metabolismo , Adenosina/inmunología , Subgrupos de Linfocitos B/inmunología , Proteínas Ligadas a GPI/inmunología , Proteínas Ligadas a GPI/metabolismo , Persona de Mediana Edad , Antígenos CD/metabolismo , Antígenos CD/inmunología , Adulto Joven , Índice de Severidad de la Enfermedad
3.
Mol Biol Rep ; 51(1): 894, 2024 Aug 08.
Artículo en Inglés | MEDLINE | ID: mdl-39115571

RESUMEN

Adenosine is a neuro- and immunomodulator that functions via G protein-coupled cell surface receptors. Several microbes, including viruses, use the adenosine signaling pathway to escape from host defense systems. Since the recent research developments in its role in health and disease, adenosine and its signaling pathway have attracted attention for targeting to treat many diseases. The therapeutic role of adenosine has been extensively studied for neurological, cardiovascular, and inflammatory disorders and bacterial pathophysiology, but published data on the role of adenosine in viral infections are lacking. Therefore, the purpose of this review article was to explain in detail the therapeutic role of adenosine signaling against viral infections, particularly COVID-19 and HIV. Several therapeutic approaches targeting A2AR-mediated pathways are in development and have shown encouraging results in decreasing the intensity of inflammatory reaction. The hypoxia-adenosinergic mechanism provides protection from inflammation-mediated tissue injury during COVID-19. A2AR expression increased remarkably in CD39 + and CD8 + T cells harvested from HIV patients in comparison to healthy subjects. A combined in vitro treatment performed by blocking PD-1 and CD39/adenosine signaling produced a synergistic outcome in restoring the CD8 + T cells funstion in HIV patients. We suggest that A2AR is an ideal target for pharmacological interventions against viral infections because it reduces inflammation, prevents disease progression, and ultimately improves patient survival.


Asunto(s)
Síndrome de Inmunodeficiencia Adquirida , Adenosina , COVID-19 , Evasión Inmune , Receptor de Adenosina A2A , SARS-CoV-2 , Transducción de Señal , Humanos , COVID-19/inmunología , COVID-19/virología , Receptor de Adenosina A2A/metabolismo , SARS-CoV-2/inmunología , SARS-CoV-2/fisiología , SARS-CoV-2/patogenicidad , Adenosina/metabolismo , Síndrome de Inmunodeficiencia Adquirida/inmunología , Síndrome de Inmunodeficiencia Adquirida/tratamiento farmacológico , Tratamiento Farmacológico de COVID-19 , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Apirasa/metabolismo , Apirasa/inmunología
4.
J Immunol ; 213(5): 588-599, 2024 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-38975728

RESUMEN

The ecto-ATPase CD39 is expressed on exhausted CD8+ T cells in chronic viral infection and has been proposed as a marker of tumor-specific CD8+ T cells in cancer, but the role of CD39 in an effector and memory T cell response has not been clearly defined. We report that CD39 is expressed on Ag-specific CD8+ short-lived effector cells, while it's co-ectoenzyme, CD73, is found on memory precursor effector cells (MPECs) in vivo. Inhibition of CD39 enzymatic activity during in vitro T cell priming enhances MPEC differentiation in vivo after transfer and infection. The enriched MPEC phenotype is associated with enhanced tissue resident memory T cell (TRM cell) establishment in the brain and salivary gland following an acute intranasal viral infection, suggesting that CD39 ATPase activity plays a role in memory CD8+ T cell differentiation. We also show that CD39 is expressed on human and murine TRM cells across several nonlymphoid tissues and melanoma, whereas CD73 is expressed on both circulating and resident memory subsets in mice. In contrast to exhausted CD39+ T cells in chronic infection, CD39+ TRM cells are fully functional when stimulated ex vivo with cognate Ag, further expanding the identity of CD39 beyond a T cell exhaustion marker.


Asunto(s)
Antígenos CD , Apirasa , Linfocitos T CD8-positivos , Diferenciación Celular , Células T de Memoria , Animales , Apirasa/inmunología , Apirasa/metabolismo , Ratones , Linfocitos T CD8-positivos/inmunología , Antígenos CD/metabolismo , Antígenos CD/inmunología , Humanos , Células T de Memoria/inmunología , Diferenciación Celular/inmunología , Memoria Inmunológica/inmunología , Ratones Endogámicos C57BL , 5'-Nucleotidasa/metabolismo , 5'-Nucleotidasa/inmunología
5.
Front Immunol ; 15: 1397967, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38947317

RESUMEN

Introduction: CD39 plays an important role in the immunoregulation and inhibition of effector cells. It is expressed on immune cells, including Tregs, and on extracellular vesicles (EVs) budding from the plasma membrane. Platelet transfusion may induce alloimmunization against HLA-I antigens, leading to refractoriness to platelet transfusion with severe consequences for patients. Tregs may play a key role in determining whether alloimmunization occurs in patients with hematologic disorders. We hypothesized that CD39+ EVs might play an immunoregulatory role, particularly in the context of platelet transfusions in patients with hematologic disorders. Such alloimmunization leads to the production of alloantibodies and is sensitive to the regulatory action of CD39. Methods: We characterized CD39+ EVs in platelet concentrates by flow cytometry. The absolute numbers and cellular origins of CD39+ EVs were evaluated. We also performed functional tests to evaluate interactions with immune cells and their functions. Results: We found that CD39+ EVs from platelet concentrates had an inhibitory phenotype that could be transferred to the immune cells with which they interacted: CD4+ and CD8+ T lymphocytes (TLs), dendritic cells, monocytes, and B lymphocytes (BLs). Moreover, the concentration of CD39+ EVs in platelet concentrates varied and was very high in 10% of concentrates. The number of these EVs present was determinant for EV-cell interactions. Finally, functional interactions were observed with BLs, CD4+ TLs and CD39+ EVs for immunoglobulin production and lymphoproliferation, with potential implications for the immunological management of patients.


Asunto(s)
Plaquetas , Vesículas Extracelulares , Tetraspanina 29 , Humanos , Vesículas Extracelulares/inmunología , Vesículas Extracelulares/metabolismo , Plaquetas/inmunología , Plaquetas/metabolismo , Tetraspanina 29/metabolismo , Comunicación Celular/inmunología , Transfusión de Plaquetas , Femenino , Linfocitos B/inmunología , Linfocitos B/metabolismo , Masculino , Apirasa/metabolismo , Apirasa/inmunología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Antígenos CD
6.
Front Immunol ; 15: 1415102, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-39007132

RESUMEN

Human regulatory T cells (Treg) suppress other immune cells. Their dysfunction contributes to the pathophysiology of autoimmune diseases, including type 1 diabetes (T1D). Infusion of Tregs is being clinically evaluated as a novel way to prevent or treat T1D. Genetic modification of Tregs, most notably through the introduction of a chimeric antigen receptor (CAR) targeting Tregs to pancreatic islets, may improve their efficacy. We evaluated CAR targeting of human Tregs to monocytes, a human ß cell line and human islet ß cells in vitro. Targeting of HLA-A2-CAR (A2-CAR) bulk Tregs to HLA-A2+ cells resulted in dichotomous cytotoxic killing of human monocytes and islet ß cells. In exploring subsets and mechanisms that may explain this pattern, we found that CD39 expression segregated CAR Treg cytotoxicity. CAR Tregs from individuals with more CD39low/- Tregs and from individuals with genetic polymorphism associated with lower CD39 expression (rs10748643) had more cytotoxicity. Isolated CD39- CAR Tregs had elevated granzyme B expression and cytotoxicity compared to the CD39+ CAR Treg subset. Genetic overexpression of CD39 in CD39low CAR Tregs reduced their cytotoxicity. Importantly, ß cells upregulated protein surface expression of PD-L1 and PD-L2 in response to A2-CAR Tregs. Blockade of PD-L1/PD-L2 increased ß cell death in A2-CAR Treg co-cultures suggesting that the PD-1/PD-L1 pathway is important in protecting islet ß cells in the setting of CAR immunotherapy. In summary, introduction of CAR can enhance biological differences in subsets of Tregs. CD39+ Tregs represent a safer choice for CAR Treg therapies targeting tissues for tolerance induction.


Asunto(s)
Apirasa , Receptores Quiméricos de Antígenos , Linfocitos T Reguladores , Humanos , Apirasa/inmunología , Apirasa/metabolismo , Linfocitos T Reguladores/inmunología , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/genética , Receptores Quiméricos de Antígenos/metabolismo , Citotoxicidad Inmunológica , Islotes Pancreáticos/inmunología , Islotes Pancreáticos/metabolismo , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/terapia , Antígeno HLA-A2/inmunología , Antígeno HLA-A2/genética , Antígeno HLA-A2/metabolismo , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Células Secretoras de Insulina/inmunología , Células Secretoras de Insulina/metabolismo , Antígenos CD
8.
Clin Immunol ; 264: 110260, 2024 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-38788885

RESUMEN

Sjögren's disease (SjD) is a chronic autoimmune disease characterized by focal lymphocytic inflammation in lacrimal and salivary glands. We recently identified IL-27 as a requisite signal for the spontaneous SjD-like manifestations in nonobese diabetic (NOD) mice. Here, we define T cell-intrinsic effects of IL-27 in lacrimal gland disease in NOD mice. IL-27 receptor was required by both CD4 T effector (Te) cells and CD8 T cells to mediate focal inflammation. Intrinsic IL-27 signaling was associated with PD-1 and ICOS expressing T follicular helper (Tfh)-like CD4 Te cells within lacrimal glands, including subsets defined by CD73 or CD39 expression. CD8 T cells capable of IL-27 signaling also expressed PD-1 with subsets expressing ICOS and CD73 demonstrating a T follicular cytotoxic (Tfc)-like cell phenotype and others expressing a CD39hi exhausted-like phenotype. These findings suggest IL-27 is a key early signal driving a follicular-type response in lacrimal gland inflammation in NOD mice.


Asunto(s)
Linfocitos T CD8-positivos , Modelos Animales de Enfermedad , Aparato Lagrimal , Ratones Endogámicos NOD , Síndrome de Sjögren , Animales , Síndrome de Sjögren/inmunología , Ratones , Linfocitos T CD8-positivos/inmunología , Aparato Lagrimal/inmunología , Aparato Lagrimal/patología , Interleucinas/inmunología , Interleucinas/metabolismo , Linfocitos T CD4-Positivos/inmunología , Receptor de Muerte Celular Programada 1/inmunología , Receptor de Muerte Celular Programada 1/metabolismo , Femenino , Transducción de Señal/inmunología , Receptores de Interleucina/inmunología , Interleucina-27/metabolismo , Interleucina-27/inmunología , Proteína Coestimuladora de Linfocitos T Inducibles/inmunología , Proteína Coestimuladora de Linfocitos T Inducibles/metabolismo , Apirasa/inmunología , Apirasa/metabolismo
9.
Cell Mol Life Sci ; 81(1): 231, 2024 May 23.
Artículo en Inglés | MEDLINE | ID: mdl-38780647

RESUMEN

CD200 is an anti-inflammatory protein that facilitates signal transduction through its receptor, CD200R, in cells, resulting in immune response suppression. This includes reducing M1-like macrophages, enhancing M2-like macrophages, inhibiting NK cell cytotoxicity, and downregulating CTL responses. Activation of CD200R has been found to modulate dendritic cells, leading to the induction or enhancement of Treg cells expressing Foxp3. However, the precise mechanisms behind this process are still unclear. Our previous study demonstrated that B cells in Peyer's patches can induce Treg cells, so-called Treg-of-B (P) cells, through STAT6 phosphorylation. This study aimed to investigate the role of CD200 in Treg-of-B (P) cell generation. To clarify the mechanisms, we used wild-type, STAT6 deficient, and IL-24 deficient T cells to generate Treg-of-B (P) cells, and antagonist antibodies (anti-CD200 and anti-IL-20RB), an agonist anti-CD200R antibody, CD39 inhibitors (ARL67156 and POM-1), a STAT6 inhibitor (AS1517499), and soluble IL-20RB were also applied. Our findings revealed that Peyer's patch B cells expressed CD200 to activate the CD200R on T cells and initiate the process of Treg-of-B (P) cells generation. CD200 and CD200R interaction triggers the phosphorylation of STAT6, which regulated the expression of CD200R, CD39, and IL-24 in T cells. CD39 regulated the expression of IL-24, which sustained the expression of CD223 and IL-10 and maintained the cell viability. In summary, the generation of Treg-of-B (P) cells by Peyer's patch B cells was through the CD200R-STAT6-CD39-IL-24 axis pathway.


Asunto(s)
Linfocitos B , Factor de Transcripción STAT6 , Linfocitos T Reguladores , Animales , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Ratones , Linfocitos B/inmunología , Linfocitos B/metabolismo , Factor de Transcripción STAT6/metabolismo , Ratones Endogámicos C57BL , Receptores de Orexina/metabolismo , Receptores de Orexina/genética , Antígenos CD/metabolismo , Antígenos CD/genética , Antígenos CD/inmunología , Transducción de Señal , Fosforilación , Ganglios Linfáticos Agregados/inmunología , Ganglios Linfáticos Agregados/metabolismo , Ganglios Linfáticos Agregados/citología , Apirasa/metabolismo , Apirasa/inmunología , Glicoproteínas de Membrana
10.
Future Oncol ; 20(19): 1367-1380, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38652041

RESUMEN

CD39 is the rate-limiting enzyme for the molecular signal cascade leading to the generation of ADP and adenosine monophosphate (AMP). In conjunction with CD73, CD39 converts adenosine triphosphate (ATP) to ADP and AMP, which leads to the accumulation of immunosuppressive adenosine in the tumor microenvironment. This review focuses on the role of CD39 and CD73 in immune response and malignant progression, including the expression of CD39 within the tumor microenvironment and its relationship to immune effector cells, and its role in antigen presentation. The role of CD39- and CD73-targeting therapeutics and cancer-directed clinical trials investigating CD39 modulation are also explored.


[Box: see text].


Asunto(s)
5'-Nucleotidasa , Apirasa , Neoplasias , Microambiente Tumoral , Humanos , 5'-Nucleotidasa/metabolismo , 5'-Nucleotidasa/inmunología , Apirasa/metabolismo , Apirasa/inmunología , Neoplasias/inmunología , Neoplasias/metabolismo , Microambiente Tumoral/inmunología , Animales , Proteínas Ligadas a GPI/inmunología , Proteínas Ligadas a GPI/metabolismo , Antígenos CD/metabolismo , Antígenos CD/inmunología , Presentación de Antígeno/inmunología
11.
PLoS Pathog ; 20(4): e1012191, 2024 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-38683845

RESUMEN

An imbalance between suppressor and effector immune responses may preclude cure in chronic parasitic diseases. In the case of Trypanosoma cruzi infection, specialized regulatory Foxp3+ T (Treg) cells suppress protective type-1 effector responses. Herein, we investigated the kinetics and underlying mechanisms behind the regulation of protective parasite-specific CD8+ T cell immunity during acute T. cruzi infection. Using the DEREG mouse model, we found that Treg cells play a role during the initial stages after T. cruzi infection, restraining the magnitude of CD8+ T cell responses and parasite control. Early Treg cell depletion increased the frequencies of polyfunctional short-lived, effector T cell subsets, without affecting memory precursor cell formation or the expression of activation, exhaustion and functional markers. In addition, Treg cell depletion during early infection minimally affected the antigen-presenting cell response but it boosted CD4+ T cell responses before the development of anti-parasite effector CD8+ T cell immunity. Crucially, the absence of CD39 expression on Treg cells significantly bolstered effector parasite-specific CD8+ T cell responses, preventing increased parasite replication in T. cruzi infected mice adoptively transferred with Treg cells. Our work underscores the crucial role of Treg cells in regulating protective anti-parasite immunity and provides evidence that CD39 expression by Treg cells represents a key immunomodulatory mechanism in this infection model.


Asunto(s)
Antígenos CD , Apirasa , Linfocitos T CD8-positivos , Enfermedad de Chagas , Linfocitos T Reguladores , Trypanosoma cruzi , Animales , Enfermedad de Chagas/inmunología , Linfocitos T Reguladores/inmunología , Linfocitos T CD8-positivos/inmunología , Ratones , Trypanosoma cruzi/inmunología , Antígenos CD/inmunología , Antígenos CD/metabolismo , Apirasa/inmunología , Apirasa/metabolismo , Ratones Endogámicos C57BL , Modelos Animales de Enfermedad
12.
J Immunother Cancer ; 10(8)2022 08.
Artículo en Inglés | MEDLINE | ID: mdl-35981785

RESUMEN

BACKGROUND: An important mechanism, by which cancer cells achieve immune escape, is the release of extracellular adenosine into their microenvironment. Adenosine activates adenosine A2A and A2B receptors on immune cells constituting one of the strongest immunosuppressive mediators. In addition, extracellular adenosine promotes angiogenesis, tumor cell proliferation, and metastasis. Cancer cells upregulate ectonucleotidases, most importantly CD39 and CD73, which catalyze the hydrolysis of extracellular ATP to AMP (CD39) and further to adenosine (CD73). Inhibition of CD39 is thus expected to be an effective strategy for the (immuno)therapy of cancer. However, suitable small molecule inhibitors for CD39 are not available. Our aim was to identify drug-like CD39 inhibitors and evaluate them in vitro. METHODS: We pursued a repurposing approach by screening a self-compiled collection of approved, mostly ATP-competitive protein kinase inhibitors, on human CD39. The best hit compound was further characterized and evaluated in various orthogonal assays and enzyme preparations, and on human immune and cancer cells. RESULTS: The tyrosine kinase inhibitor ceritinib, a potent anticancer drug used for the treatment of anaplastic lymphoma kinase (ALK)-positive metastatic non-small cell lung cancer, was found to strongly inhibit CD39 showing selectivity versus other ectonucleotidases. The drug displays a non-competitive, allosteric mechanism of CD39 inhibition exhibiting potency in the low micromolar range, which is independent of substrate (ATP) concentration. We could show that ceritinib inhibits ATP dephosphorylation in peripheral blood mononuclear cells in a dose-dependent manner, resulting in a significant increase in ATP concentrations and preventing adenosine formation from ATP. Importantly, ceritinib (1-10 µM) substantially inhibited ATP hydrolysis in triple negative breast cancer and melanoma cells with high native expression of CD39. CONCLUSIONS: CD39 inhibition might contribute to the effects of the powerful anticancer drug ceritinib. Ceritinib is a novel CD39 inhibitor with high metabolic stability and optimized physicochemical properties; according to our knowledge, it is the first brain-permeant CD39 inhibitor. Our discovery will provide the basis (i) to develop more potent and balanced dual CD39/ALK inhibitors, and (ii) to optimize the ceritinib scaffold towards interaction with CD39 to obtain potent and selective drug-like CD39 inhibitors for future in vivo studies.


Asunto(s)
Antineoplásicos , Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Adenosina/metabolismo , Adenosina Trifosfato/metabolismo , Antígenos CD/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Apirasa/inmunología , Apirasa/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Humanos , Inmunoterapia , Leucocitos Mononucleares/metabolismo , Neoplasias Pulmonares/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinas , Proteínas Tirosina Quinasas Receptoras , Sulfonas , Microambiente Tumoral
13.
Front Immunol ; 13: 946698, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35967385

RESUMEN

Multiple sclerosis (MS) is a highly disabling, progressive neurodegenerative disease with no curative treatment available. Although significant progress has been made in understanding how MS develops, there remain aspects of disease pathogenesis that are yet to be fully elucidated. In this regard, studies have shown that dysfunctional adenosinergic signaling plays a pivotal role, as patients with MS have altered levels adenosine (ADO), adenosine receptors and proteins involved in the generation and termination of ADO signaling, such as CD39 and adenosine deaminase (ADA). We have therefore performed a literature review regarding the involvement of the adenosinergic system in the development of MS and propose mechanisms by which the modulation of this system can support drug development and repurposing.


Asunto(s)
Esclerosis Múltiple , Enfermedades Neurodegenerativas , Receptores Purinérgicos P1 , Adenosina/inmunología , Adenosina Desaminasa/inmunología , Apirasa/inmunología , Humanos , Esclerosis Múltiple/etiología , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/terapia , Enfermedades Neurodegenerativas/etiología , Enfermedades Neurodegenerativas/inmunología , Enfermedades Neurodegenerativas/terapia , Receptores Purinérgicos P1/inmunología , Transducción de Señal
14.
PLoS One ; 17(7): e0264566, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35901034

RESUMEN

Current medical guidelines consider pregnant women with COVID-19 to be a high-risk group. Since physiological gestation downregulates the immunological response to maintain "maternal-fetal tolerance", SARS-CoV-2 infection may constitute a potentially threatening condition to both the mother and the fetus. To establish the immune profile in pregnant COVID-19+ patients, a cross-sectional study was conducted. Pregnant women with COVID-19 (P-COVID-19+; n = 15) were analyzed and compared with nonpregnant women with COVID-19 (NP-COVID-19+; n = 15) or those with physiological pregnancy (P-COVID-19-; n = 13). Serological cytokine and chemokine concentrations, leucocyte immunophenotypes, and mononuclear leucocyte responses to polyclonal stimuli were analyzed in all groups. Higher concentrations of serological TNF-α, IL-6, MIP1b and IL-4 were observed within the P-COVID-19+ group, while cytokines and chemokines secreted by peripheral leucocytes in response to LPS, IL-6 or PMA-ionomicin were similar among the groups. Immunophenotype analysis showed a lower percentage of HLA-DR+ monocytes in P-COVID-19+ than in P-COVID-19- and a higher percentage of CD39+ monocytes in P-COVID-19+ than in NP-COVID-19+. After whole blood polyclonal stimulation, similar percentages of T cells and TNF+ monocytes between groups were observed. Our results suggest that P-COVID-19+ elicits a strong inflammatory response similar to NP-COVID19+ but also displays an anti-inflammatory response that controls the ATP/adenosine balance and prevents hyperinflammatory damage in COVID-19.


Asunto(s)
COVID-19 , Monocitos , Apirasa/inmunología , Estudios Transversales , Citocinas , Femenino , Humanos , Interleucina-6 , Embarazo , SARS-CoV-2
15.
Gastroenterology ; 163(4): 965-981.e31, 2022 10.
Artículo en Inglés | MEDLINE | ID: mdl-35738329

RESUMEN

BACKGROUND & AIMS: Exhaustion of CD8 T cells has been suggested to inform different clinical outcomes in Crohn's disease, but detailed analyses are lacking. This study aimed to identify the role of exhaustion on a single-cell level and identify relevant CD8 T cell populations in Crohn's disease. METHODS: Blood and intestinal tissue from 58 patients with Crohn's disease (active disease or remission) were assessed for CD8 T cell expression of exhaustion markers and their cytokine profile by highly multiplexed flow and mass cytometry. Key disease-associated subsets were sorted and analyzed by RNA sequencing. CD39 inhibition assays were performed in vitro. RESULTS: Activated CD39+ and CD39+PD-1+ CD8 T cell subsets expressing multiple exhaustion markers were enriched at low frequency in active Crohn's disease. Their cytokine production capacity was inversely linked to the Harvey-Bradshaw Index. Subset-level protein and transcriptome profiling revealed co-existence of effector and exhaustion programs in CD39+ and CD39+ PD-1+CD8 T cells, with CD39+ cells likely originating from the intestine. CD39 enzymatic activity controlled T cell cytokine production. Importantly, transcriptional exhaustion signatures were enriched in remission in CD39-expressing subsets with up-regulation of TOX. Subset-level transcriptomics revealed a CD39-related gene module that is associated with the clinical course. CONCLUSIONS: These data showed a role for the exhaustion of peripheral CD39-expressing CD8 T cell subsets in Crohn's disease. Their low frequency illustrated the utility of single-cell cytometry methods for identification of relevant immune populations. Importantly, the link of their exhaustion status to the clinical activity and their specific gene signatures have implications for exhaustion-based personalized medicine approaches.


Asunto(s)
Apirasa , Linfocitos T CD8-positivos , Enfermedad de Crohn , Apirasa/sangre , Apirasa/genética , Apirasa/inmunología , Biomarcadores/sangre , Linfocitos T CD8-positivos/inmunología , Enfermedad de Crohn/sangre , Enfermedad de Crohn/genética , Enfermedad de Crohn/inmunología , Citocinas/inmunología , Humanos , Pronóstico , Receptor de Muerte Celular Programada 1/genética , Receptor de Muerte Celular Programada 1/inmunología , Subgrupos de Linfocitos T
16.
J Immunother Cancer ; 10(6)2022 06.
Artículo en Inglés | MEDLINE | ID: mdl-35688560

RESUMEN

BACKGROUND: Adjuvant immune checkpoint inhibitor (ICI) immunotherapies have significantly reduced the recurrence rate in high-risk patients with stage III melanoma compared with surgery alone. However, 48% of anti-PD-1-treated patients will develop recurrent disease within 4 years. There is a need to identify biomarkers of recurrence after adjuvant ICI to enable identification of patients in need of alternative treatment strategies. As cytotoxic T cells are critical for the antitumor response to anti-PD-1, we sought to determine whether specific subsets were predictive of recurrence in anti-PD-1-treated high-risk patients with stage III melanoma. METHODS: Associations with recurrence in patients with stage III melanoma were sought by analyzing resection specimens (n=103) taken prior to adjuvant nivolumab/pembrolizumab±low-dose/low-interval ipilimumab. Multiplex immunohistochemistry was used to quantify intratumoral CD8+ T-cell populations using phenotypical markers CD39, CD103, and PD-1. RESULTS: With a median follow-up of 19.3 months, 37/103 (36%) of patients had a recurrence. Two CD8+ T-cell subpopulations were significantly associated with recurrence. First, CD39+ tumor-resident memory cells (CD39+CD103+PD-1+CD8+ (CD39+ Trm)) comprised a significantly higher proportion of CD8+ T cells in recurrence-free patients (p=0.0004). Conversely, bystander T cells (CD39-CD103-PD-1-CD8+) comprised a significantly greater proportion of T cells in patients who developed recurrence (p=0.0002). Spatial analysis identified that CD39+ Trms localized significantly closer to melanoma cells than bystander T cells. Multivariable analysis confirmed significantly improved recurrence-free survival (RFS) in patients with a high proportion of intratumoral CD39+ Trms (1-year RFS high 78.1% vs low 49.9%, HR 0.32, 95% CI 0.15 to 0.69), no complete lymph node dissection performed, and less advanced disease stage (HR 2.85, 95% CI 1.13 to 7.19, and HR 1.29, 95% CI 0.59 to 2.82). The final Cox regression model identified patients who developed recurrence with an area under the curve of 75.9% in the discovery cohort and 69.5% in a separate validation cohort (n=33) to predict recurrence status at 1 year. CONCLUSIONS: Adjuvant immunotherapy-treated patients with a high proportion of CD39+ Trms in their baseline melanoma resection have a significantly reduced risk of melanoma recurrence. This population of T cells may not only represent a biomarker of RFS following anti-PD-1 therapy, but may also be an avenue for therapeutic manipulation and enhancing outcomes for immunotherapy-treated patients with cancer.


Asunto(s)
Melanoma , Neoplasias Cutáneas , Apirasa/inmunología , Humanos , Inmunoterapia , Melanoma/tratamiento farmacológico , Receptor de Muerte Celular Programada 1/uso terapéutico , Neoplasias Cutáneas/patología , Melanoma Cutáneo Maligno
17.
Cell Mol Life Sci ; 79(6): 298, 2022 May 18.
Artículo en Inglés | MEDLINE | ID: mdl-35585332

RESUMEN

BACKGROUND: Mood disorders have been associated with risk of clinical relapses in multiple sclerosis (MS), a demyelinating disease mediated by myelin-specific T cells. OBJECTIVES: We aimed to investigate the impact of major depressive disorder (MDD) and cytokine profile of T-cells in relapsing remitting MS patients. METHODS: For our study, plasma and PBMC were obtained from 60 MS patients (30 with lifetime MDD) in remission phase. The PBMC cultures were stimulated with anti-CD3/anti-CD28 beads or myelin basic protein (MBP), and effector and regulatory T cell phenotypes were determined by flow cytometry. The cytokine levels, both in the plasma or in the supernatants collected from PBMC cultures, were quantified by Luminex. In some experiments, the effect of serotonin (5-HT) was investigated. RESULTS: Here, higher Th17-related cytokine levels in response to anti-CD3/anti-CD28 and MBP were quantified in the plasma and PBMC cultures of the MS/MDD group in comparison with MS patients. Further, elevated frequency of CD4+ and CD8+ T cells capable of producing IL-17, IL-22 and GM-CSF was observed in depressed patients. Interestingly, the percentage of myelin-specific IFN-γ+IL-17+ and IFN-γ+GM-CSF+ CD4+ T cells directly correlated with neurological disabilities. In contrast, the occurrence of MDD reduced the proportion of MBP-specific CD39+Tregs subsets. Notably, the severity of both neurological disorder and depressive symptoms inversely correlated with these Tregs. Finally, the addition of 5-HT downregulated the release of Th17-related cytokines in response to anti-CD3/anti-CD28 and myelin antigen. CONCLUSIONS: In summary, our findings suggested that recurrent major depression, by favoring imbalances of effector Th17 and Treg cell subsets, contributes to MS severity.


Asunto(s)
Apirasa , Autoantígenos , Trastorno Depresivo Mayor , Esclerosis Múltiple , Vaina de Mielina , Linfocitos T Reguladores , Células Th17 , Apirasa/inmunología , Autoantígenos/inmunología , Linfocitos T CD8-positivos/inmunología , Citocinas/inmunología , Trastorno Depresivo Mayor/sangre , Trastorno Depresivo Mayor/diagnóstico , Trastorno Depresivo Mayor/inmunología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/inmunología , Humanos , Interleucina-17/inmunología , Leucocitos Mononucleares/inmunología , Esclerosis Múltiple/inmunología , Vaina de Mielina/inmunología , Serotonina/inmunología , Linfocitos T Reguladores/inmunología , Células Th17/inmunología
18.
J Med Virol ; 94(5): 2089-2101, 2022 05.
Artículo en Inglés | MEDLINE | ID: mdl-35032133

RESUMEN

COVID-19 is a disease characterized by acute respiratory failure and is a major health problem worldwide. Here, we aimed to investigate the role of CD39 expression in Treg cell subsets in COVID-19 immunopathogenesis and its relationship to disease severity. One hundred and ninety COVID-19 patients (juveniles, adults) and 43 volunteers as healthy controls were enrolled in our study. Flow cytometric analysis was performed using a 10-color monoclonal antibody panel from peripheral blood samples. In adult patients, CD39+ Tregs increased with disease severity. In contrast, CD39+ Tregs were decreased in juvenile patients in an age-dependent manner. Overall, our study reveals an interesting profile of CD39-expressing Tregs in adult and juvenile cases of COVID-19. Our results provide a better understanding of the possible role of Tregs in the mechanism of immune response in COVID-19 cases.


Asunto(s)
Apirasa , COVID-19 , Linfocitos T Reguladores , Adulto , Apirasa/biosíntesis , Apirasa/inmunología , Apirasa/metabolismo , COVID-19/inmunología , COVID-19/metabolismo , Factores de Transcripción Forkhead , Humanos , Índice de Severidad de la Enfermedad , Subgrupos de Linfocitos T/inmunología , Linfocitos T Reguladores/inmunología
19.
Eur J Immunol ; 52(1): 96-108, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34505280

RESUMEN

The detection of tumor-specific T cells in solid tumors is integral to interrogate endogenous antitumor responses and to advance downstream therapeutic applications. Multiple biomarkers are reported to identify endogenous tumor-specific tumor-infiltrating lymphocytes (TILs), namely CD137, PD-1, CD103, and CD39; however, a direct comparison of these molecules has yet to be performed. We evaluated these biomarkers in primary human ovarian tumor samples using single-cell mass cytometry to compare their relative phenotypic profiles, and examined their response to autologous tumor cells ex vivo. PD-1+ , CD103+ , and CD39+ TILs all contain a CD137+ cell subset, while CD137+ TILs highly co-express the aforementioned markers. CD137+ TILs exhibit the highest expression of cytotoxic effector molecules compared to PD-1+ , CD103+ , or CD39+ TILs. Removal of CD137+ cells from PD-1+ , CD103+ , or CD39+ TILs diminish their IFN-γ secretion in response to autologous tumor cell stimulation, while CD137+ TILs maintain high HLA-dependent IFN-γ secretion. CD137+ TILs exhibited an exhausted phenotype but with CD28 co-expression, suggesting possible receptiveness to reinvigoration via immune checkpoint blockade. Together, our findings demonstrate that the antitumor abilities of PD-1+ , CD103+ , and CD39+ TILs are mainly derived from a subset of CD137-expressing TILs, implicating CD137 as a more selective biomarker for naturally occurring tumor-specific TILs.


Asunto(s)
Antígenos CD/inmunología , Apirasa/inmunología , Biomarcadores de Tumor/inmunología , Cadenas alfa de Integrinas/inmunología , Linfocitos Infiltrantes de Tumor/inmunología , Neoplasias Ováricas/inmunología , Receptor de Muerte Celular Programada 1/inmunología , Miembro 9 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/inmunología , Femenino , Humanos , Interferón gamma/inmunología
20.
Comput Math Methods Med ; 2021: 2725799, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34790251

RESUMEN

Type 1 regulatory T (Tr1) cells play a fundamental role in maintaining and inducing immune tolerance. Our preliminary study demonstrated that an interleukin- (IL-) 10-mediated pathway is a possible regulatory mechanism underlying the xenoantigen-specific human Treg enhanced suppressive capacity. Here, we developed a feasible protocol for expanding IL-10-induced xenoantigen-specific human Tr1 cells in vitro which would be more efficient in transplantation immunotherapy efficiency. In this study, xenoantigen-specific Tr1 cells are generated from human naive CD4+ T cells expanded for two subsequent xenoantigen-stimulation cycles with recombinant human IL-10. The phenotype and suppressive capacity of xenoantigen-stimulated Tr1 cells are assessed, and the mechanism of their suppression is studied. Tr1 cells can be induced by porcine xenoantigen stimulation combined with IL-10, IL-2, and IL-15, displaying an increased expression of CD49b, CTLA-4, and LAG-3 without expressing Foxp3 which also showed an effector memory Treg phenotype and expressed high levels of CD39. After xenoantigen stimulation, the IL-10 and IL-5 gene expression in Tr1 cells increased, secreting more IL-10, and xenoantigen-stimulated Tr1 cells changed their T cell receptor (TCR) Vß repertoire, increasing the expression of TCR Vß2, TCR Vß9, and TCR Vß13. In a pig to human mixed lymphocyte reaction (MLR), xenoantigen-stimulated Tr1 cells displayed enhanced suppressive capacity via CD39 in a dose-dependent manner. Moreover, IL-5 could affect the proliferation of xenoantigen-specific Tr1 cells, but not their phenotypes' expression. This study provides a theory and feasible method for immune tolerance induction in clinical xenotransplantation.


Asunto(s)
Antígenos Heterófilos/administración & dosificación , Tolerancia Inmunológica , Linfocitos T Reguladores/inmunología , Traslado Adoptivo , Adulto , Animales , Apirasa/inmunología , Proliferación Celular , Técnicas de Cocultivo , Biología Computacional , Femenino , Humanos , Técnicas In Vitro , Interleucina-10/biosíntesis , Interleucina-5/antagonistas & inhibidores , Interleucina-5/inmunología , Activación de Linfocitos , Masculino , Persona de Mediana Edad , Sus scrofa , Linfocitos T Reguladores/clasificación , Linfocitos T Reguladores/citología , Inmunología del Trasplante , Tolerancia al Trasplante
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