RESUMEN
Innate pattern recognition receptor agonists, including Toll-like receptors (TLRs), alter the tumor microenvironment and prime adaptive antitumor immunity. However, TLR agonists present toxicities associated with widespread immune activation after systemic administration. To design a TLR-based therapeutic suitable for systemic delivery and capable of safely eliciting tumor-targeted responses, we developed immune-stimulating antibody conjugates (ISACs) comprising a TLR7/8 dual agonist conjugated to tumor-targeting antibodies. Systemically administered human epidermal growth factor receptor 2 (HER2)-targeted ISACs were well tolerated and triggered a localized immune response in the tumor microenvironment that resulted in tumor clearance and immunological memory. Mechanistically, ISACs required tumor antigen recognition, Fcγ-receptor-dependent phagocytosis and TLR-mediated activation to drive tumor killing by myeloid cells and subsequent T-cell-mediated antitumor immunity. ISAC-mediated immunological memory was not limited to the HER2 ISAC target antigen since ISAC-treated mice were protected from rechallenge with the HER2- parental tumor. These results provide a strong rationale for the clinical development of ISACs.
Asunto(s)
Inmunoterapia , Neoplasias , Inmunidad Adaptativa , Animales , Antígenos de Neoplasias , Inmunoterapia/métodos , Ratones , Neoplasias/tratamiento farmacológico , Microambiente TumoralRESUMEN
Dendritic cells (DCs) are powerful antigen presenting cells, derived from bone marrow progenitors (cDCs) and monocytes (moDCs), that can shape the immune response by priming either proinflammatory or tolerogenic immune effector cells. The cellular mechanisms responsible for the generation of DCs that will prime a proinflammatory or tolerogenic response are poorly understood. Here we describe a novel mechanism by which tolerogenic DCs are formed from monocytes. When human monocytes were cultured with CD4+FoxP3+ natural regulatory T cells (Tregs) and T helper cells (Th) from healthy donor blood, they differentiated into regulatory DCs (DC Reg ), capable of generating induced Tregs from naïve T cells. DC Reg exhibited morphology, surface phenotype, cytokine secretion, and transcriptome that were distinct from other moDCs including those derived from monocytes cultured with Th or with GM-CSF/IL-4, as well as macrophages (MΦ). Direct cell contact between monocytes, Tregs and Th, along with Treg-derived CTLA-4, IL-10 and TGF-ß, was required for the phenotypic differentiation of DC Reg , although only IL-10 was required for imprinting the Treg-inducing capacity of DC Reg . High ratios of Treg:Th, along with monocytes and DC Reg similar in function and phenotype to those induced in vitro, were present in situ in human colorectal cancer specimens. Thus, through the combined actions of Tregs and Th, monocytes differentiate into DCs with regulatory properties, forming a positive feedback loop to reinforce Treg initiated immune regulation. This mechanism may contribute to immune tolerance in tissues such as tumors, which contain an abundance of Tregs, Th and monocytes.
Asunto(s)
Comunicación Celular , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Monocitos/inmunología , Transducción de Señal , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Biomarcadores , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Comunicación Celular/inmunología , Diferenciación Celular , Neoplasias del Colon/inmunología , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Biología Computacional/métodos , Perfilación de la Expresión Génica , Humanos , Inmunomodulación , Inmunofenotipificación , Monocitos/metabolismo , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , TranscriptomaRESUMEN
BM-derived DC (BMDC) are powerful antigen-presenting cells. When loaded with immune complexes (IC), consisting of tumor antigens bound to antitumor antibody, BMDC induce powerful antitumor immunity in mice. However, attempts to employ this strategy clinically with either tumor-associated DC (TADC) or monocyte-derived DC (MoDC) have been disappointing. To investigate the basis for this phenomenon, we compared the response of BMDC, TADC, and MoDC to tumor IgG-IC. Our findings revealed, in both mice and humans, that upon exposure to IgG-IC, BMDC internalized the IC, increased costimulatory molecule expression, and stimulated autologous T cells. In contrast, TADC and, surprisingly, MoDC remained inert upon contact with IC due to dysfunctional signaling following engagement of Fcγ receptors. Such dysfunction is associated with elevated levels of the Src homology region 2 domain-containing phosphatase-1 (SHP-1) and phosphatases regulating Akt activation. Indeed, concomitant inhibition of both SHP-1 and phosphatases that regulate Akt activation conferred upon TADC and MoDC the capacity to take up and process IC and induce antitumor immunity in vivo. This work identifies the molecular checkpoints that govern activation of MoDC and TADC and their capacity to elicit T cell immunity.
Asunto(s)
Complejo Antígeno-Anticuerpo/inmunología , Antígenos de Neoplasias/inmunología , Células Dendríticas/inmunología , Neoplasias Pulmonares/inmunología , Proteína Tirosina Fosfatasa no Receptora Tipo 6/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Diferenciación Celular , Línea Celular Tumoral , Humanos , Activación de Linfocitos , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/citología , Recurrencia Local de Neoplasia , Neoplasias Experimentales/inmunología , Linfocitos T/inmunología , Microambiente TumoralRESUMEN
Type 2 diabetes mellitus (T2D) is a metabolic disease that is strongly tied to obesity and often preceded by insulin resistance (IR). It has been established that chronic inflammation of hypertrophic adipose tissue depots in obese individuals leads to obesity-associated IR and is mediated by cells of the innate immune system, particularly macrophages. More recently, cells of the adaptive immune system, B and T lymphocytes, have also emerged as important regulators of glucose homeostasis, raising the intriguing possibility that antigen-driven immune responses play a role in disease. In this review, we critically evaluate the roles that various B and T cell subsets play in IR, and then we examine the data suggesting that antigen-driven mechanisms, such as antigen presentation and costimulation, may drive the activity of these lymphocytes.
Asunto(s)
Inmunidad Adaptativa , Resistencia a la Insulina , Obesidad/metabolismo , Animales , Presentación de Antígeno , Linfocitos B/inmunología , Antígeno B7-1/fisiología , Antígeno B7-2/fisiología , Humanos , Memoria Inmunológica , Linfocitos T/inmunologíaRESUMEN
Obesity-associated insulin resistance, a common precursor of type 2 diabetes, is characterized by chronic inflammation of tissues, including visceral adipose tissue (VAT). Here we show that B-1a cells, a subpopulation of B lymphocytes, are novel and important regulators of this process. B-1a cells are reduced in frequency in obese high-fat diet (HFD)-fed mice, and EGFP interleukin-10 (IL-10) reporter mice show marked reductions in anti-inflammatory IL-10 production by B cells in vivo during obesity. In VAT, B-1a cells are the dominant producers of B cell-derived IL-10, contributing nearly half of the expressed IL-10 in vivo. Adoptive transfer of B-1a cells into HFD-fed B cell-deficient mice rapidly improves insulin resistance and glucose tolerance through IL-10 and polyclonal IgM-dependent mechanisms, whereas transfer of B-2 cells worsens metabolic disease. Genetic knockdown of B cell-activating factor (BAFF) in HFD-fed mice or treatment with a B-2 cell-depleting, B-1a cell-sparing anti-BAFF antibody attenuates insulin resistance. These findings establish B-1a cells as a new class of immune regulators that maintain metabolic homeostasis and suggest manipulation of these cells as a potential therapy for insulin resistance.
Asunto(s)
Subgrupos de Linfocitos B/fisiología , Resistencia a la Insulina/fisiología , Animales , Glucemia , Grasas de la Dieta/administración & dosificación , Grasas de la Dieta/toxicidad , Femenino , Regulación de la Expresión Génica/fisiología , Intolerancia a la Glucosa , Humanos , Inmunoglobulina M , Interleucina-10/genética , Interleucina-10/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Obesidad/inducido químicamente , Obesidad/metabolismoRESUMEN
Organ transplant recipients face life-long immunosuppression and consequently are at high risk of comorbidities. Occasionally, kidney transplant recipients develop a state of targeted immune quiescence (operational tolerance) against an HLA-mismatched graft, allowing them to withdraw all immunosuppression and retain stable graft function while resuming immune responses to third-party antigens. Methods to better understand and monitor this state of alloimmune quiescence by transcriptional profiling may reveal a gene signature that identifies patients for whom immunosuppression could be titrated to reduce patient and graft morbidities. Therefore, we investigated 571 unique peripheral blood samples from 348 HLA-mismatched renal transplant recipients and 101 nontransplant controls in a four-stage study including microarray, quantitative PCR, and flow cytometry analyses. We report a refined and highly validated (area under the curve, 0.95; 95% confidence interval, 0.92 to 0.97) peripheral blood three-gene assay (KLF6, BNC2, CYP1B1) to detect the state of operational tolerance by quantitative PCR. The frequency of predicted alloimmune quiescence in stable renal transplant patients receiving long-term immunosuppression (n=150) was 7.3% by the three-gene assay. Targeted cell sorting of peripheral blood from operationally tolerant patients showed a significant shift in the ratio of circulating monocyte-derived dendritic cells with significantly different expression of the genes constituting the three-gene assay. Our results suggest that incorporation of patient screening by specific cellular and gene expression assays may support the safety of drug minimization trials and protocols.
Asunto(s)
Biomarcadores/sangre , Terapia de Inmunosupresión , Trasplante de Riñón , Inmunología del Trasplante/genética , Adolescente , Adulto , Recuento de Células Sanguíneas , Antígeno CD11c/metabolismo , Estudios de Casos y Controles , Niño , Citocromo P-450 CYP1B1/genética , Citocromo P-450 CYP1B1/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Células Dendríticas/metabolismo , Femenino , Perfilación de la Expresión Génica , Humanos , Factor 6 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Adulto JovenRESUMEN
Monocytes rapidly infiltrate inflamed tissues and differentiate into CD209(+) inflammatory dendritic cells (DCs) that promote robust immunity or, if unregulated, inflammatory disease. Previous studies in experimental animal models indicate that inflammatory DC depletion through systemic elimination of their monocyte precursors with clodronate-loaded liposomes ameliorates the development of psoriasis and other diseases. However, translation of systemic monocyte depletion strategies is difficult due to the importance of monocytes during homeostasis and infection clearance. Here, we describe a strategy that avoids the monocyte intermediates to deplete inflammatory DCs through antibody-loaded toxin. Mice with an abundance of inflammatory DCs as a consequence of lipopolysaccharide exposure were treated with anti-CD209 antibody conjugated to saporin, a potent ribosome inactivator. The results demonstrate depletion of CD209(+) DCs. This strategy could prove useful for the targeted reduction of inflammatory DCs in disease.
Asunto(s)
Anticuerpos Monoclonales/inmunología , Moléculas de Adhesión Celular/antagonistas & inhibidores , Células Dendríticas/inmunología , Inmunotoxinas/inmunología , Lectinas Tipo C/antagonistas & inhibidores , Procedimientos de Reducción del Leucocitos , Receptores de Superficie Celular/antagonistas & inhibidores , Proteínas Inactivadoras de Ribosomas Tipo 1/inmunología , Animales , Moléculas de Adhesión Celular/metabolismo , Células Dendríticas/metabolismo , Modelos Animales de Enfermedad , Citometría de Flujo , Inflamación/inmunología , Inflamación/metabolismo , Lectinas Tipo C/metabolismo , Ratones , Receptores de Superficie Celular/metabolismo , SaporinasRESUMEN
Two critical functions of dendritic cells (DC) are to activate and functionally polarize T cells. Activated T cells can, in turn, influence DC maturation, although their effect on de novo DC development is poorly understood. Here we report that activation of T cells in mice, with either an anti-CD3 antibody or super antigen, drives the rapid formation of CD209(+)CD11b(+)CD11c(+) MHC II(+) DC from monocytic precursors (Mo-DC). GM-CSF is produced by T cells following activation, but surprisingly, it is not required for the formation of CD209(+) Mo-DC. CD40L, however, is critical for the full induction of Mo-DC following T cell activation. T cell induced CD209(+) Mo-DC are comparable to conventional CD209(-) DC in their ability to stimulate T cell proliferation. However, in contrast to conventional CD209(-) DC, CD209(+) Mo-DC fail to effectively polarize T cells, as indicated by a paucity of T cell cytokine production. The inability of CD209(+) Mo-DC to polarize T cells is partly explained by increased expression of PDL-2, since blockade of this molecule restores some polarizing capacity to the Mo-DC. These findings expand the range of signals capable of driving Mo-DC differentiation in vivo beyond exogenous microbial factors to include endogenous factors produced following T cell activation.
Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Células Dendríticas/citología , Células Dendríticas/metabolismo , Lectinas Tipo C/metabolismo , Activación de Linfocitos/inmunología , Proteína 2 Ligando de Muerte Celular Programada 1/metabolismo , Receptores de Superficie Celular/metabolismo , Subgrupos de Linfocitos T/inmunología , Animales , Ligando de CD40/metabolismo , Diferenciación Celular/inmunología , Movimiento Celular , Células Dendríticas/inmunología , Femenino , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Ganglios Linfáticos/inmunología , Ratones , Ratones Noqueados , Monocitos/inmunología , Subgrupos de Linfocitos T/metabolismoRESUMEN
In chronically inflamed tissues, such as those affected by autoimmune disease, activated Th cells often colocalize with monocytes. We investigate in this study how murine Th cells influence the phenotype and function of monocytes. The data demonstrate that Th1, Th2, and Th17 subsets promote the differentiation of autologous monocytes into MHC class II(+), CD11b(+), CD11c(+) DC that we call DCTh. Although all Th subsets induce the formation of DCTh, activated Th17 cells uniquely promote the formation of IL-12/IL-23-producing DCTh (DCTh17) that can polarize both naive and Th17 cells to a Th1 phenotype. In the inflamed CNS of mice with Th17-mediated experimental autoimmune encephalomyelitis, Th cells colocalize with DC, as well as monocytes, and the Th cells obtained from these lesions drive the formation of DCTh that are phenotypically indistinguishable from DCTh17 and polarize naive T cells toward a Th1 phenotype. These results suggest that DCTh17 are critical in the interplay of Th17- and Th1-mediated responses and may explain the previous finding that IL-17-secreting Th cells become IFN-γ-secreting Th1 cells in experimental autoimmune encephalomyelitis and other autoimmune disorders.
Asunto(s)
Autoinmunidad/inmunología , Células Dendríticas/inmunología , Inflamación/inmunología , Células TH1/inmunología , Células Th17/inmunología , Animales , Antígeno CD11b/metabolismo , Antígeno CD11c/metabolismo , Diferenciación Celular , Movimiento Celular , Polaridad Celular , Células Cultivadas , Sistema Nervioso Central/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Femenino , Antígenos de Histocompatibilidad Clase II/metabolismo , Interferón gamma/metabolismo , Interleucina-12/biosíntesis , Interleucina-17/metabolismo , Interleucina-23/biosíntesis , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/inmunologíaRESUMEN
Cytokines dimerize their receptors, with the binding of the 'second chain' triggering signaling. In the interleukin (IL)-4 and IL-13 system, different cell types express varying numbers of alternative second receptor chains (γc or IL-13Rα1), forming functionally distinct type I or type II complexes. We manipulated the affinity and specificity of second chain recruitment by human IL-4. A type I receptor-selective IL-4 'superkine' with 3,700-fold higher affinity for γc was three- to ten-fold more potent than wild-type IL-4. Conversely, a variant with high affinity for IL-13Rα1 more potently activated cells expressing the type II receptor and induced differentiation of dendritic cells from monocytes, implicating the type II receptor in this process. Superkines showed signaling advantages on cells with lower second chain numbers. Comparative transcriptional analysis reveals that the superkines induce largely redundant gene expression profiles. Variable second chain numbers can be exploited to redirect cytokines toward distinct cell subsets and elicit new actions, potentially improving the selectivity of cytokine therapy.
Asunto(s)
Citocinas/fisiología , Interleucina-4/análogos & derivados , Interleucina-4/farmacología , Linfocitos T CD4-Positivos/metabolismo , Diferenciación Celular/efectos de los fármacos , Línea Celular , Células Dendríticas/efectos de los fármacos , Citometría de Flujo , Perfilación de la Expresión Génica , Humanos , Factores Inmunológicos/farmacología , Interleucina-4/química , Mutación/fisiología , Fenotipo , Fosforilación , Ingeniería de Proteínas , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Interleucina-4/efectos de los fármacos , Factor de Transcripción STAT6/metabolismo , Transducción de Señal/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
Monocytes and T helper (T(H)) cells rapidly infiltrate inflamed tissues where monocytes differentiate into inflammatory dendritic cells (DCs) through undefined mechanisms. Our studies indicate that T(H) cells frequently interact with monocytes in inflamed skin and elicit the differentiation of specialized DC subsets characteristic of these lesions. In psoriasis lesions, T(H)1 and T(H)17 cells interact with monocytes and instruct these cells to differentiate into T(H)1- and T(H)17-promoting DCs, respectively. Correspondingly, in acute atopic dermatitis, T(H)2 cells interact with monocytes and elicit the formation of T(H)2-promoting DCs. DC formation requires GM-CSF and cell contact, whereas T(H) subset specific cytokines dictate DC function and the expression of DC subset specific surface molecules. Moreover, the phenotypes of T cell-induced DC subsets are maintained after subsequent stimulation with a panel of TLR agonists, suggesting that T(H)-derived signals outweigh downstream TLR signals in their influence on DC function. These findings indicate that T(H) cells govern the formation and function of specialized DC subsets.
Asunto(s)
Citocinas/biosíntesis , Células Dendríticas/inmunología , Monocitos/inmunología , Psoriasis/metabolismo , Transducción de Señal , Piel/metabolismo , Células TH1/inmunología , Células Th17/inmunología , Células Th2/inmunología , Antígenos CD/análisis , Antígenos CD/biosíntesis , Comunicación Celular/inmunología , Diferenciación Celular/inmunología , Técnicas de Cocultivo , Citocinas/inmunología , Células Dendríticas/citología , Células Dendríticas/metabolismo , Citometría de Flujo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/inmunología , Humanos , Inmunohistoquímica , Inmunofenotipificación , Activación de Linfocitos , Monocitos/citología , Monocitos/metabolismo , Psoriasis/inmunología , Psoriasis/patología , Transducción de Señal/inmunología , Piel/inmunología , Piel/patología , Células TH1/citología , Células TH1/metabolismo , Células Th17/citología , Células Th17/metabolismo , Células Th2/citología , Células Th2/metabolismo , Receptores Toll-Like/inmunologíaRESUMEN
Chronic inflammation characterized by T cell and macrophage infiltration of visceral adipose tissue (VAT) is a hallmark of obesity-associated insulin resistance and glucose intolerance. Here we show a fundamental pathogenic role for B cells in the development of these metabolic abnormalities. B cells accumulate in VAT in diet-induced obese (DIO) mice, and DIO mice lacking B cells are protected from disease despite weight gain. B cell effects on glucose metabolism are mechanistically linked to the activation of proinflammatory macrophages and T cells and to the production of pathogenic IgG antibodies. Treatment with a B cell-depleting CD20 antibody attenuates disease, whereas transfer of IgG from DIO mice rapidly induces insulin resistance and glucose intolerance. Moreover, insulin resistance in obese humans is associated with a unique profile of IgG autoantibodies. These results establish the importance of B cells and adaptive immunity in insulin resistance and suggest new diagnostic and therapeutic modalities for managing the disease.
Asunto(s)
Linfocitos B/inmunología , Inmunoglobulina G/biosíntesis , Resistencia a la Insulina/inmunología , Linfocitos T/inmunología , Animales , Autoanticuerpos/biosíntesis , Autoantígenos/inmunología , Autoinmunidad , Grasas de la Dieta/efectos adversos , Glucosa/metabolismo , Humanos , Cadenas mu de Inmunoglobulina/genética , Inflamación/inmunología , Grasa Intraabdominal/inmunología , Activación de Linfocitos , Depleción Linfocítica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/etiología , Obesidad/inmunología , Obesidad/metabolismoRESUMEN
Interleukin (IL)-9-producing CD4(+) T cells are a novel subset of T helper (Th) cells that develops independently of the Th1, Th2, Th17 and regulatory T-cell lineages. Similar to the murine model, transforming growth factor (TGF)-beta and IL-4 directed human naive CD4(+) T cells to produce IL-9. Whereas IL-4 suppressed TGF-beta-induced Foxp3 expression, TGF-beta failed to inhibit IL-4-mediated upregulation of the Th2 transcription factor GATA-3. Addition of IL-1 beta, IL-6, IL-10, interferon (IFN)-alpha, IFN-beta or IL-21 to Th9-polarizing conditions augmented Th9 differentiation, while the Th1-associated cytokines IFN-gamma and IL-27 partially suppressed IL-9 production. Given that T cells are a primary source of IL-21, IL-21 expression was analyzed under Th9-polarizing conditions in the context of inflammatory cytokines. Surprisingly, type I IFNs induced elevated levels of IL-21, and blockade of IL-21 abrogated their ability to enhance Th9 differentiation. Taken together, these data indicate a complex cytokine network in the regulation of human IL-9-producing CD4(+) T cells.