RESUMO
Odronextamab is a fully-human IgG4-based CD20xCD3 bispecific antibody that binds to CD3 on T cells and CD20 on B cells, triggering T-cell-mediated cytotoxicity independent of T-cell-receptor recognition. Adequate safety, tolerability, and encouraging durable complete responses have been observed in an ongoing first-in-human (FIH) study of odronextamab in patients with relapsed/refractory (R/R) B-cell non-Hodgkin lymphoma (B-NHL; NCT02290951). We retrospectively evaluated the pharmacokinetic, pharmacodynamic, and antitumor characteristics of odronextamab in a series of in vitro/in vivo preclinical experiments, to assess their translational value to inform dose escalation for the FIH study. Half-maximal effective concentration values from in vitro cytokine release assays (range: 0.05-0.08 mg/L) provided a reasonable estimate of odronextamab concentrations in patients associated with cytokine release at a 0.5 mg dose (maximum serum concentration: 0.081 mg/L) on week 1/day 1, which could therefore be used to determine the week 1 clinical dose. Odronextamab concentrations resulting in 100% inhibition of tumor growth in a Raji xenograft tumor mouse model (1-10 mg/L) were useful to predict efficacious concentrations in patients and inform dose-escalation strategy. Although predicted human pharmacokinetic parameters derived from monkey data overestimated projected odronextamab exposure, they provided a conservative estimate for FIH starting doses. With step-up dosing, the highest-tested weekly odronextamab dose in patients (320 mg) exceeded the 1 mg/kg single dose in monkeys without step-up dosing. In conclusion, combination of odronextamab in vitro cytokine data, efficacious concentration data from mouse tumor models, and pharmacokinetic evaluations in monkeys has translational value to inform odronextamab FIH study design in patients with R/R B-NHL.
Assuntos
Anticorpos Biespecíficos , Antineoplásicos , Linfoma de Células B , Animais , Antígenos CD20 , Antineoplásicos/uso terapêutico , Citocinas , Humanos , Linfoma de Células B/tratamento farmacológico , Linfoma de Células B/patologia , Camundongos , Estudos RetrospectivosRESUMO
NK cells have been shown to either promote or protect from autoimmune diseases. Several studies have examined the role of receptors preferentially expressed by NK cells in the spontaneous disease of NOD mice or the direct role of NK cells in acute induced disease models of diabetes. Yet, the role of NK cells in spontaneous diabetes has not been directly addressed. Here, we used the NOD.NK1.1 congenic mouse model to examine the role of NK cells in spontaneous diabetes. Significant numbers of NK cells were only seen in the pancreas of mice with disease. Pancreatic NK cells displayed an activated surface phenotype and proliferated more than NK cells from other tissues in the diseased mice. Nonetheless, depletion of NK cells had no effect on dendritic cell maturation or T cell proliferation. In spontaneous disease, the deletion of NK cells had no significant impact on disease onset. NK cells were also not required to promote disease induced by adoptively transferred pathogenic CD4(+) T cells. Thus, NK cells are not required for spontaneous autoimmune diabetes in NOD mice.
Assuntos
Diabetes Mellitus Tipo 1/imunologia , Células Matadoras Naturais/imunologia , Animais , Diferenciação Celular/imunologia , Membrana Celular/metabolismo , Proliferação de Células , Citocinas/metabolismo , Citotoxicidade Imunológica , Células Dendríticas/imunologia , Progressão da Doença , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/patologia , Fígado/imunologia , Fígado/patologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos NOD , Pâncreas/imunologia , Pâncreas/patologia , FenótipoRESUMO
OBJECTIVE: The progressive infiltration of pancreatic islets by lymphocytes is mandatory for development of autoimmune type 1 diabetes. This inflammatory process is mediated by several mediators that are potential therapeutic targets to arrest development of type 1 diabetes. In this study, we investigate the role of one of these mediators, interleukin-16 (IL-16), in the pathogenesis of type 1 diabetes in NOD mice. RESEARCH DESIGN AND METHODS: At different stages of progression of type 1 diabetes, we characterized IL-16 in islets using GEArray technology and immunoblot analysis and also quantitated IL-16 activity in cell migration assays. IL-16 expression was localized in islets by immunofluorescence and confocal imaging. In vivo neutralization studies were performed to assess the role of IL-16 in the pathogenesis of type 1 diabetes. RESULTS: The increased expression of IL-16 in islets correlated with the development of invasive insulitis. IL-16 immunoreactivity was found in islet infiltrating T-cells, B-cells, NK-cells, and dendritic cells, and within an insulitic lesion, IL-16 was derived from infiltrating cells. CD4(+) and CD8(+) T-cells as well as B220(+) B-cells were identified as sources of secreted IL-16. Blockade of IL-16 in vivo protected against type 1 diabetes by interfering with recruitment of CD4(+) T-cells to the pancreas, and this protection required the activity of the chemokine CCL4. CONCLUSIONS: IL-16 production by leukocytes in islets augments the severity of insulitis during the onset of type 1 diabetes. IL-16 and CCL4 appear to function as counterregulatory proteins during disease development. Neutralization of IL-16 may represent a novel therapy for the prevention of type 1 diabetes.
Assuntos
Tetracloreto de Carbono/farmacologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Interleucina-16/fisiologia , Transferência Adotiva , Animais , Linfócitos T CD4-Positivos/imunologia , Movimento Celular , Diabetes Mellitus Tipo 1/fisiopatologia , Hiperinsulinismo/fisiopatologia , Immunoblotting , Interleucina-16/genética , Ilhotas Pancreáticas/imunologia , Ilhotas Pancreáticas/fisiologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Pâncreas/fisiologia , Pâncreas/fisiopatologia , RNA Mensageiro/genética , Linfócitos T/imunologiaRESUMO
Sjögren's Syndrome (SS) is a human autoimmune disease characterized by immune-mediated destruction of the lacrimal and salivary glands. In this study, we show that the Aire-deficient mouse represents a new tool to investigate autoimmune dacryoadenitis and keratoconjunctivitis sicca, features of SS. Previous work in the Aire-deficient mouse suggested a role for alpha-fodrin, a ubiquitous Ag, in the disease process. Using an unbiased biochemical approach, however, we have identified a novel lacrimal gland autoantigen, odorant binding protein 1a, targeted by the autoimmune response. This novel autoantigen is expressed in the thymus in an Aire-dependent manner. The results from our study suggest that defects in central tolerance may contribute to SS and provide a new and clinically relevant model to investigate the pathogenic mechanisms in lacrimal gland autoimmunity and associated ocular surface sequelae.
Assuntos
Autoanticorpos/biossíntese , Síndromes do Olho Seco/genética , Síndromes do Olho Seco/imunologia , Receptores Odorantes/imunologia , Síndrome de Sjogren/genética , Síndrome de Sjogren/imunologia , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , Animais , Autoanticorpos/sangue , Dacriocistite/genética , Dacriocistite/imunologia , Dacriocistite/patologia , Modelos Animais de Doenças , Síndromes do Olho Seco/patologia , Feminino , Humanos , Ceratoconjuntivite Seca/genética , Ceratoconjuntivite Seca/imunologia , Ceratoconjuntivite Seca/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Nus , Camundongos SCID , Receptores Odorantes/biossíntese , Receptores Odorantes/genética , Síndrome de Sjogren/patologia , Timo/imunologia , Timo/metabolismo , Timo/patologia , Proteína AIRERESUMO
Autoimmune pancreatitis (AIP) is a heterogeneous autoimmune disease in humans characterized by a progressive lymphocytic and plasmacytic infiltrate in the exocrine pancreas. In this study, we report that regulatory T cell-deficient NOD.CD28KO mice spontaneously develop AIP that closely resembles the human disease. NOD mouse AIP was associated with severe periductal and parenchymal inflammation of the exocrine pancreas by CD4(+) T cells, CD8(+) T cells, and B cells. Spleen CD4(+) T cells were found to be both necessary and sufficient for the development of AIP. Autoantibodies and autoreactive T cells from affected mice recognized a approximately 50-kDa protein identified as pancreatic amylase. Importantly, administration of tolerogenic amylase-coupled fixed spleen cells significantly ameliorated disease severity, suggesting that this protein functions as a key autoantigen. The establishment and characterization of this spontaneous pancreatic amylase-specific AIP in regulatory T cell-deficient NOD.CD28KO mice provides an excellent model for the study of disease pathogenesis and development of new therapies for human autoimmune pancreatitis.
Assuntos
Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Antígenos CD28/genética , Pancreatite/genética , Pancreatite/imunologia , Transferência Adotiva , Animais , Autoantígenos/imunologia , Doenças Autoimunes/enzimologia , Doenças Autoimunes/terapia , Antígenos CD28/biossíntese , Células Cultivadas , Feminino , Tolerância Imunológica/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Transgênicos , Pâncreas/enzimologia , Pâncreas/imunologia , Pâncreas/patologia , Pancreatite/enzimologia , Pancreatite/terapia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Linfócitos T Reguladores/transplante , alfa-Amilases/imunologia , alfa-Amilases/metabolismoRESUMO
We previously reported that interleukin (IL)-4 treatment of nonobese diabetic (NOD) mice elevates intrapancreatic CCL4 expression and protects from type 1 diabetes. Here, we show that antibody neutralization of CCL4 abrogates the ability of T-cells from IL-4-treated NOD mice to transfer protection against type 1 diabetes. Intradermal delivery of CCL4 via a plasmid vector stabilized by incorporation of the Epstein-Barr virus EBNA1/oriP episomal maintenance replicon (pHERO8100-CCL4) to NOD mice beginning at later stages of disease progression protects against type 1 diabetes. This protection was associated with a Th2-like response in the spleen and pancreas; decreased recruitment of activated CD8(+) T-cells to islets, accompanied by diminished CCR5 expression on CD8(+) T-cells; and regulatory T-cell activity in the draining pancreatic lymph nodes. Thus, inflammatory responses that target islet beta-cells are suppressed by CCL4, which implicates the use of CCL4 therapeutically to prevent type 1 diabetes.
Assuntos
Quimiocinas CC/metabolismo , Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 1/prevenção & controle , Células Secretoras de Insulina/patologia , Envelhecimento , Animais , Quimiocina CCL4 , Quimiocinas CC/genética , Diabetes Mellitus Tipo 1/metabolismo , Terapia Genética , Inflamação/prevenção & controle , Interleucina-4/imunologia , Interleucina-4/farmacologia , Transplante das Ilhotas Pancreáticas , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Baço/citologia , Baço/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T/transplanteRESUMO
Deficiencies in NKT cell number and function mediate the development of Type 1 diabetes (TID). NKT cell activation with the CD1d ligand alpha-galactosylceramide (alpha-GalCer) corrects these deficiencies and prevents the onset and recurrence of T1D in NOD mice. To investigate how alpha-GalCer accomplishes this, we conducted three sets of studies. First, gene microarray analyses showed that alpha-GalCer treatment decreases interleukin (IL)16 and increases IL10 and MIP1beta gene expression in the spleen. Anti-IL16 antibody treatment protects NOD mice against insulitis and T1D, and neutralization of MIP1beta abrogates IL4 induced protection from T1D. Second, alpha-GalCer treatment of NOD.ILA(-/-) mice demonstrated that IL4 expression is required for prevention of T1D but not for NKT cell development. Third, we found that diabetes resistance in three novel congenic NOD.B6Idd4 mouse strains is associated with an increased number of NKT cells in pancreatic lymph nodes (PLNs). This increase was not evident in the spleen or PLNs of NOD.MIP1a(-/-) mice after alpha-GalCer treatment. Our data suggest that MIP1beta is a candidate gene in Idd4 that regulates NKT cell function and diabetes susceptibility. By controlling the expression and activity of IL16 and MIP1beta alpha-GalCer treatment may modulate the number, localization and function of NKT cells and regulate susceptibility to T1D.
Assuntos
Antígenos CD1/imunologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Células Matadoras Naturais/imunologia , Transferência Adotiva , Animais , Antígenos CD1d , Quimiocinas/genética , Diabetes Mellitus Tipo 1/genética , Regulação da Expressão Gênica/imunologia , Humanos , Camundongos , Camundongos Endogâmicos NOD , Receptores de Quimiocinas/genéticaRESUMO
Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is expressed in different tissues and cells, including pancreas and lymphocytes, and can induce apoptosis in various tumor cells but not in most normal cells. The specific roles of TRAIL in health and disease remain unclear. Here we show by cDNA array analyses that TRAIL gene expression is upregulated in pancreatic islets during the development of autoimmune type 1 diabetes in nonobese diabetic (NOD) mice and in Min6 islet beta-cells activated by TNF-alpha + interferon-gamma. However, stimulation of freshly isolated pancreatic islets or Min6 cells with TRAIL did not induce their apoptosis. TRAIL blockade exacerbates the onset of type 1 diabetes in NOD.Scid recipients of transferred diabetogenic T-cells and in cyclophosphamide-treated NOD mice. TRAIL inhibits the proliferation of NOD diabetogenic T-cells by suppressing interleukin (IL)-2 production and cell cycle progression, and this inhibition can be rescued in the presence of exogenous IL-2. cDNA array and Western blot analyses indicate that TRAIL upregulates the expression of the cdk inhibitor p27(kip1). Our data suggest that TRAIL is an important immune regulator of the development of type 1 diabetes.
Assuntos
Apoptose/imunologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/fisiopatologia , Glicoproteínas de Membrana/genética , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Animais , Proteínas Reguladoras de Apoptose , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Divisão Celular/imunologia , Linhagem Celular , Inibidor de Quinase Dependente de Ciclina p27 , Diabetes Mellitus Tipo 1/patologia , Feminino , Ilhotas Pancreáticas/patologia , Ilhotas Pancreáticas/fisiologia , Ligantes , Glicoproteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Análise de Sequência com Séries de Oligonucleotídeos , Linfócitos T/citologia , Ligante Indutor de Apoptose Relacionado a TNF , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismoRESUMO
Activation-induced cell death (AICD) is a mechanism of homeostasis that limits the clonal expansion of autoreactive T cells and regulates central and peripheral tolerance. In nonobese diabetic (NOD) mice, defects in central and peripheral tolerance are associated with a proliferative hyporesponsiveness of thymocytes and peripheral T cells elicited upon TCR activation. We investigated whether these defects in tolerance induction and hyporesponsiveness of NOD T cells manifest in an altered susceptibility to TCR-induced AICD. TCR-activated NOD splenic CD4+ and CD8+ T cells are more resistant to AICD than control strain C57BL/6, BALB/c, and NOR T cells. NOR CD4+ but not CD8+ T cells are resistant to TCR-induced AICD. Whereas c-FLIP expression is reduced in activated T cells from control strains, it persists in activated NOD CD8+ T cells and is accompanied by diminished activity of caspase-3 and -8. IL-4 reduces this c-FLIP expression and increases caspase-3 and -8 activity in activated NOD CD8+ T cells. Moreover, IL-4 and CD28 costimulation restores the susceptibility of NOD CD8+ T cells to AICD, and this is associated with increased expression of CD25, CD95, CD95L, and TNFR2. Thus, deficient activation of CD8+ T cells and their greater resistance to TCR-induced AICD may mediate defective peripheral tolerance and the development of T1D in NOD mice.
Assuntos
Apoptose/imunologia , Linfócitos T CD8-Positivos/imunologia , Diabetes Mellitus Tipo 1/imunologia , Tolerância Imunológica/imunologia , Peptídeos e Proteínas de Sinalização Intracelular , Animais , Antígenos CD/imunologia , Antígenos CD/metabolismo , Proteína Reguladora de Apoptosis Semelhante a CASP8 e FADD , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Proteínas de Transporte/antagonistas & inibidores , Proteínas de Transporte/biossíntese , Caspase 3 , Caspase 8 , Caspase 9 , Caspases/imunologia , Caspases/metabolismo , Ativação Enzimática , Feminino , Interleucina-2/farmacologia , Interleucina-4/imunologia , Interleucina-4/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Receptores de Antígenos de Linfócitos T/imunologia , Organismos Livres de Patógenos EspecíficosRESUMO
Twenty diabetes susceptibility loci on 12 mouse chromosomes have been identified to control the development of type 1 diabetes at the level of either initiation of insulitis or progression from insulitis to overt diabetes or both. Previously, we demonstrated that the genetic control of T-cell proliferative unresponsiveness in nonobese diabetic (NOD) mice is linked to Idd4 on mouse chromosome 11. Here, we show by congenic mapping of three newly generated NOD.B6Idd4 diabetes-resistant mouse strains that Idd4 is limited to a 5.2-cM interval of chromosome 11. This B6-derived region expressed in NOD.B6Idd4A mice maps between the D11Nds1 (43.8 cM) and D11Mit38/D11Mit325 (49.0 cM) markers and dramatically reduces the development of both insulitis and type 1 diabetes. NOD.B6Idd4B and NOD.B6Idd4C mice, which carry a smaller B6-derived segment of chromosome 11 that spans <5.2 cM distal to D11Nds1, exhibit protection against type 1 diabetes with the restoration of T-cell proliferation. Our findings suggest that diabetes resistance conferred by Idd4 may be mediated by the Idd4.1 and Idd4.2 subloci. Idd4.1 is localized in the D11Nds1 interval that influences both diabetes and insulitis. Idd4.2 is localized within the D11Mit38/325 interval that mainly influences diabetes incidence and restores T-cell proliferative responsiveness. Three potential candidate genes, platelet activating factor acetylhydrolase Ib1, nitric oxide synthase-2, and CC chemokine genes, are localized in the 5.2-cM interval.