RESUMO
The ability of immune therapies to control cancer has recently generated intense interest. This therapeutic outcome is reliant on T cell recognition of tumour cells. The natural function of dendritic cells (DC) is to generate adaptive responses, by presenting antigen to T cells, hence they are a logical target to generate specific anti-tumour immunity. Our understanding of the biology of DC is expanding, and they are now known to be a family of related subsets with variable features and function. Most clinical experience to date with DC vaccination has been using monocyte-derived DC vaccines. There is now growing experience with alternative blood-derived DC derived vaccines, as well as with multiple forms of tumour antigen and its loading, a wide range of adjuvants and different modes of vaccine delivery. Key insights from pre-clinical studies, and lessons learned from early clinical testing drive progress towards improved vaccines. The potential to fortify responses with other modalities of immunotherapy makes clinically effective "second generation" DC vaccination strategies a priority for cancer immune therapists.
Assuntos
Células Dendríticas/imunologia , Células Dendríticas/transplante , Imunoterapia , Neoplasias/imunologia , Neoplasias/terapia , Humanos , Linfócitos T/imunologiaRESUMO
Previous studies demonstrated that endogenous glucocorticoid signaling in osteoblasts promotes inflammation in murine immune arthritis. The current study determined whether disruption of endogenous glucocorticoid signaling in chondrocytes also modulates the course and severity of arthritis. Tamoxifen-inducible chondrocyte-targeted glucocorticoid receptor-knockout (chGRKO) mice were generated by breeding GRflox/flox mice with tamoxifen-inducible collagen 2a1 Cre (Col2a1-CreERT2) mice. Antigen-induced arthritis (AIA) and K/BxN serum transfer-induced arthritis (STIA) were induced in both chGRKO mice and their Cre-negative GRflox/flox littermates [wild type (WT)]. Arthritis was assessed by measurement of joint swelling and histology of joints collected at d 14. Neutrophil activity and gene expression patterns associated with cartilage damage were also evaluated. In both arthritis models clinical (joint swelling) and histologic indices of inflammatory activity were significantly greater in chGRKO than in WT mice. The STIA model was characterized by early up-regulation of CXCR2/CXCR2 ligand gene expression in ankle tissues, and significant and selective expansion of splenic CXCR2+ neutrophils in chGRKO arthritic compared to WT arthritic mice. At later stages, gene expression of enzymes involved in cartilage degradation was up-regulated in chGRKO but not WT arthritic mice. Therefore, we summarize that chondrocytes actively mitigate local joint inflammation, cartilage degradation and systemic neutrophil activity via a glucocorticoid-dependent pathway.-Tu, J., Stoner, S., Fromm, P. D., Wang, T., Chen, D., Tuckermann, J., Cooper, M. S., Seibel, M. J., Zhou, H. Endogenous glucocorticoid signaling in chondrocytes attenuates joint inflammation and damage.
Assuntos
Artrite Experimental/metabolismo , Condrócitos/metabolismo , Glucocorticoides/metabolismo , Animais , Artrite Experimental/etiologia , Artrite Experimental/patologia , Osso e Ossos/metabolismo , Osso e Ossos/patologia , Quimiocinas/genética , Quimiocinas/metabolismo , Mediadores da Inflamação/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Transgênicos , Receptores de Glucocorticoides/deficiência , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Transdução de Sinais , Regulação para CimaRESUMO
Chemotherapy and hematopoietic stem cell transplantation are effective treatments for most Hodgkin lymphoma patients, however there remains a need for better tumor-specific target therapy in Hodgkin lymphoma patients with refractory or relapsed disease. Herein, we demonstrate that membrane CD83 is a diagnostic and therapeutic target, highly expressed in Hodgkin lymphoma cell lines and Hodgkin and Reed-Sternberg cells in 29/35 (82.9%) Hodgkin lymphoma patient lymph node biopsies. CD83 from Hodgkin lymphoma tumor cells was able to trogocytose to surrounding T cells and, interestingly, the trogocytosing CD83+T cells expressed significantly more programmed death-1 compared to CD83-T cells. Hodgkin lymphoma tumor cells secreted soluble CD83 that inhibited T-cell proliferation, and anti-CD83 antibody partially reversed the inhibitory effect. High levels of soluble CD83 were detected in Hodgkin lymphoma patient sera, which returned to normal in patients who had good clinical responses to chemotherapy confirmed by positron emission tomography scans. We generated a human anti-human CD83 antibody, 3C12C, and its toxin monomethyl auristatin E conjugate, that killed CD83 positive Hodgkin lymphoma cells but not CD83 negative cells. The 3C12C antibody was tested in dose escalation studies in non-human primates. No toxicity was observed, but there was evidence of CD83 positive target cell depletion. These data establish CD83 as a potential biomarker and therapeutic target in Hodgkin lymphoma.
Assuntos
Antígenos CD/sangue , Biomarcadores Tumorais/sangue , Doença de Hodgkin/tratamento farmacológico , Imunoglobulinas/sangue , Glicoproteínas de Membrana/sangue , Terapia de Alvo Molecular/métodos , Adolescente , Adulto , Idoso , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Antígenos CD/imunologia , Feminino , Doença de Hodgkin/diagnóstico , Humanos , Imunoglobulinas/imunologia , Masculino , Glicoproteínas de Membrana/imunologia , Pessoa de Meia-Idade , Terapia de Salvação/métodos , Linfócitos T/citologia , Adulto Jovem , Antígeno CD83RESUMO
C-type lectin receptors play important roles in immune cell interactions with the environment. We described CD302 as the simplest, single domain, type I C-type lectin receptor and showed it was expressed mainly on the myeloid phagocytes in human blood. CD302 colocalized with podosomes and lamellopodia structures, so we hypothesized that it played a role in cell adhesion or migration. In this study, we used mouse models to obtain further insights into CD302 expression and its potential immunological function. Mouse CD302 transcripts were, as in humans, highest in the liver, followed by lungs, lymph nodes (LN), spleen, and bone marrow. In liver, CD302 was expressed by hepatocytes, liver sinusoidal endothelial cells, and Kupffer cells. A detailed analysis of CD302 transcription in mouse immune cells revealed highest expression by myeloid cells, particularly macrophages, granulocytes, and myeloid dendritic cells (mDC). Interestingly, 2.5-fold more CD302 was found in migratory compared with resident mDC populations and higher CD302 expression in mouse M1 versus M2 macrophages was also noteworthy. CD302 knockout (CD302KO) mice were generated. Studies on the relevant immune cell populations revealed a decrease in the frequency and numbers of migratory mDC within CD302KO LN compared with wild-type LN. In vitro studies showed CD302KO and wild-type DC had an equivalent capacity to undergo maturation, prime T cells, uptake Ags, and migrate toward the CCL19/CCL21 chemokines. Nevertheless, CD302KO migratory DC exhibited reduced in vivo migration into LN, confirming a functional role for CD302 in mDC migration.
Assuntos
Quimiotaxia de Leucócito/fisiologia , Células Dendríticas/metabolismo , Lectinas Tipo C/metabolismo , Receptores de Superfície Celular/metabolismo , Animais , Linhagem Celular , Separação Celular , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Reação em Cadeia da PolimeraseRESUMO
CD83 is a member of the Ig gene superfamily, first identified in activated lymphocytes. Since then, CD83 has become an important marker for defining activated human dendritic cells (DC). Several potential CD83 mRNA isoforms have been described, including a soluble form detected in human serum, which may have an immunosuppressive function. To further understand the biology of CD83, we examined its expression in different human immune cell types before and after activation using a panel of mouse and human anti-human CD83 mAb. The mouse anti-human CD83 mAbs, HB15a and HB15e, and the human anti-human CD83 mAb, 3C12C, were selected to examine cytoplasmic and surface CD83 expression, based on their different binding characteristics. Glycosylation of CD83, the CD83 mRNA isoforms, and soluble CD83 released differed among blood DC, monocytes, and monocyte-derived DC, and other immune cell types. A small T cell population expressing surface CD83 was identified upon T cell stimulation and during allogeneic MLR. This subpopulation appeared specifically during viral Ag challenge. We did not observe human CD83 on unstimulated human natural regulatory T cells (Treg), in contrast to reports describing expression of CD83 on mouse Treg. CD83 expression was increased on CD4+, CD8+ T, and Treg cells in association with clinical acute graft-versus-host disease in allogeneic hematopoietic cell transplant recipients. The differential expression and function of CD83 on human immune cells reveal potential new roles for this molecule as a target of therapeutic manipulation in transplantation, inflammation, and autoimmune diseases.
Assuntos
Antígenos CD/metabolismo , Células Dendríticas/imunologia , Doença Enxerto-Hospedeiro/imunologia , Transplante de Células-Tronco Hematopoéticas , Imunoglobulinas/metabolismo , Glicoproteínas de Membrana/metabolismo , Monócitos/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Doença Aguda , Animais , Antígenos CD/genética , Antígenos Virais/imunologia , Células Cultivadas , Glicosilação , Humanos , Imunoglobulinas/genética , Ativação Linfocitária , Glicoproteínas de Membrana/genética , Camundongos , Isoformas de RNA/genética , RNA Mensageiro/genética , Transplante Homólogo , Antígeno CD83RESUMO
Although the earliestrudimentaryattempts at exploiting the immune system for cancer therapy can be traced back to the late 18th Century, it was not until the past decade that cancer immunotherapeutics have truly entered mainstream clinical practice. Given their potential to stimulate both adaptive and innate antitumor immune responses, dendritic cells (DCs) have come under intense scrutiny in recent years as pharmacological tools for cancer immunotherapy. Conceptually, the clinical effectiveness of this form of active immunotherapy relies on the completion of three critical steps: 1) the DCs used as immunotherapeutic vehicles must properly activate the antitumor immune effector cells of the host, 2) these immune effector cells must be receptive to stimulation by the DCs and be competent to mediate their antitumor effects, which 3) requires overcoming the various immune-inhibitory mechanisms used by the tumor cells. In this review, following a brief overview of the pivotal milestones in the history of cancer immunotherapy, we will introduce the reader to the basic immunobiological and pharmacological principles of active cancer immunotherapy using DCs. We will then discuss how current research is trying to define the optimal parameters for each of the above steps to realize the full clinical potential of DC therapeutics. Given its high suitability for immune interventions, acute myeloid leukemia was chosen here to showcase the latest research trends driving the field of DC-based cancer immunotherapy.
Assuntos
Células Dendríticas/metabolismo , Imunoterapia Ativa/métodos , Neoplasias/imunologia , Neoplasias/terapia , Linfócitos T Citotóxicos/imunologia , Transferência Adotiva/métodos , Anticorpos Monoclonais , Antígenos de Neoplasias/imunologia , Apoptose , Vacinas Anticâncer/imunologia , Técnicas de Cultura de Células , Citocinas/biossíntese , Células Dendríticas/imunologia , Relação Dose-Resposta a Droga , Vias de Administração de Medicamentos , Humanos , Células Matadoras Naturais/imunologia , Complexo Principal de Histocompatibilidade/imunologia , Transdução de SinaisRESUMO
The CC-chemokine receptor 6 (CCR6) can be detected on naive and activated B cells. Counterintuitively, its absence accelerates the appearance of germinal centres (GCs) and increases the production of low-affinity antibodies. The detailed mechanism of CCR6 function during the humoral response has remained elusive, but previously we identified a distinct CCR6high B-cell population in vivo early after antigenic challenge. In this study, we defined this population specifically as early, activated pre-GC B cells. In accordance, we show that CCR6 is upregulated rapidly within hours on the protein or mRNA level after activation in vitro. In addition, only activated B cells migrated specifically towards CCL20, the specific ligand for CCR6. Lack of CCR6 increased the dark zone/light zone ratio of GC and led to decreased antigen-specific IgG1 and IgG2a antibody generation in a B-cell intrinsic manner in mixed bone marrow chimeras. In contrast, antigen-specific IgM responses were normal. Hence, CCR6 negatively regulates entry of activated, antigen-specific pre-GC B cells into the GC reaction.
Assuntos
Formação de Anticorpos/imunologia , Linfócitos B/metabolismo , Centro Germinativo/metabolismo , Receptores CCR6/metabolismo , Animais , Formação de Anticorpos/efeitos dos fármacos , Linfócitos B/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Quimiocina CCL20/farmacologia , Citometria de Fluxo , Centro Germinativo/efeitos dos fármacos , Cinética , Ativação Linfocitária/efeitos dos fármacos , Camundongos Endogâmicos C57BL , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores CCR6/genética , Regulação para Cima/efeitos dos fármacosRESUMO
Human plasmacytoid dendritic cells (pDCs) were considered to be a phenotypically and functionally homogeneous cell population; however, recent analyses indicate potential heterogeneity. This is of major interest, given their importance in the induction of anti-viral responses and their role in creating immunologically permissive environments for human malignancies. For this reason, we investigated the possible presence of human pDC subsets in blood and bone marrow, using unbiased cell phenotype clustering and functional studies. This defined two major functionally distinct human pDC subsets, distinguished by differential expression of CD2. The CD2(hi) and CD2(lo) pDCs represent discontinuous subsets, each with hallmark pDC functionality, including interferon-alpha production. The rarer CD2(hi) pDC subset demonstrated a significant survival advantage over CD2(lo) pDC during stress and upon exposure to glucocorticoids (GCs), which was associated with higher expression of the anti-apoptotic molecule BCL2. The differential sensitivity of these two human pDC subsets to GCs is demonstrated in vivo by a relative increase in CD2(hi) pDC in multiple myeloma patients treated with GCs. Hence, the selective apoptosis of CD2(lo) pDC during stress represents a novel mechanism for the control of innate responses.
Assuntos
Antígenos CD2/metabolismo , Células Dendríticas/metabolismo , Estresse Fisiológico , Apoptose/efeitos dos fármacos , Medula Óssea/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Glucocorticoides/farmacologia , Humanos , Ligantes , Linfonodos/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Receptores Imunológicos/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Receptores Toll-Like/metabolismoRESUMO
The transfer of membrane proteins between cells during contact, known as trogocytosis, can create novel cells with a unique phenotype and altered function. We demonstrate that trogocytosis is more common in multiple myeloma (MM) than chronic lymphocytic leukemia and Waldenstrom macroglobulinaemia; that T cells are more probable to be recipients than B or natural killer cells; that trogocytosis occurs independently of either the T-cell receptor or HLA compatibility; and that after trogocytosis, T cells with acquired antigens can become novel regulators of T-cell proliferation. We screened 168 patients with MM and found that CD86 and human leukocyte antigen G (HLA-G) were antigens commonly acquired by T cells from malignant plasma cells. CD3+ CD86acq+ and CD3+ HLA-Gacq+ cells were more prevalent in bone marrow than peripheral blood samples. The presence of either CD86 or HLA-G on malignant plasma cells was associated with a poor prognosis. CD38++ side population cells expressed HLA-G, suggesting that these putative myeloma stem cells could generate immune tolerance. HLA-G+ T cells had a regulatory potency similar to natural Tregs, thus providing another novel mechanism for MM to avoid effective immune surveillance.
Assuntos
Antígeno B7-2/imunologia , Antígenos HLA-G/imunologia , Leucemia Linfocítica Crônica de Células B/metabolismo , Mieloma Múltiplo/metabolismo , Plasmócitos/imunologia , Macroglobulinemia de Waldenstrom/metabolismo , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biomarcadores/análise , Membrana Celular/imunologia , Membrana Celular/metabolismo , Proliferação de Células , Humanos , Tolerância Imunológica , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/mortalidade , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/mortalidade , Especificidade de Órgãos , Plasmócitos/metabolismo , Prognóstico , Transporte Proteico/imunologia , Taxa de Sobrevida , Linfócitos T/imunologia , Linfócitos T/metabolismo , Macroglobulinemia de Waldenstrom/imunologia , Macroglobulinemia de Waldenstrom/mortalidadeRESUMO
In the absence of TNF, the normally resistant C57BL/6 (B6.WT) strain develops a fatal, progressive form of leishmaniasis after infection with Leishmania major. It is not yet understood which TNF activity or the lack thereof is responsible for the dramatic progression of leishmaniasis in TNF-negative (B6.TNF(-/-)) mice. To elucidate the underlying mechanisms resulting in the fatal outcome of L. major infection in this gene-deficient mouse strain, we analyzed the monocytic component of the inflammatory infiltrate in the draining popliteal lymph node and the site of the infection using multicolor flow cytometry. The leukocytic infiltrate within the draining lymph node and footpad of B6.TNF(-/-) mice resembled that of B6.WT mice over the first 2 wk of cutaneous L. major infection. Thereafter, the B6.TNF(-/-) mice showed an increase of CD11c(+)Ly-6C(+)CCR2(+) monocytic dendritic cells within the popliteal lymph node in comparison with B6.WT mice. This increase of inflammatory dendritic cells was paired with the accumulation of a novel CD11b(+)Ly-6C(low)CCR2(low) population that was not present in B6.WT mice. This B6.TNF(-/-)- and B6.TNFR1(-/-)-specific cell population was CD115(+)Ly-6G(-)iNOS(-), not apoptotic, and harbored large numbers of parasites.
Assuntos
Leishmaniose Cutânea/imunologia , Macrófagos/imunologia , Macrófagos/parasitologia , Fator de Necrose Tumoral alfa/imunologia , Animais , Antígenos Ly/imunologia , Modelos Animais de Doenças , Citometria de Fluxo , Separação Imunomagnética , Leishmania major/imunologia , Linfonodos/citologia , Linfonodos/imunologia , Linfonodos/parasitologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Confocal , FenótipoRESUMO
The CC-chemokine receptor 6 (CCR6) is expressed constitutively at an intermediate level on naïve B cells and is upregulated after activation on pregerminal center (GC) B cells. We hypothesized that it could be involved in the events leading to GC reaction and high-affinity antibody production, and therefore investigated the potential role of CCR6 in B-cell differentiation in vivo. After antigenic challenge of CCR6-/- mice with the T-cell-dependent antigen nitrophenyl-keyhole limpet hemocyanin (NP-KLH), GC B-cell development was found to be accelerated and the number of GC had increased significantly compared with control mice, but the antibodies produced by CCR6-/- B cells were on average of lower affinity. We conclude from these data that the CCR6/CCL20 axis has an important role in regulating the kinetics and efficiency of the GC reaction.
Assuntos
Linfócitos B/imunologia , Centro Germinativo/imunologia , Receptores CCR6/metabolismo , Animais , Afinidade de Anticorpos/genética , Formação de Anticorpos/genética , Quimiocina CCL20/metabolismo , Regulação da Expressão Gênica , Centro Germinativo/citologia , Haptenos , Hemocianinas/imunologia , Imunomodulação , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores CCR6/genética , Receptores CCR6/imunologia , Regulação para CimaRESUMO
Leishmania (L.) major is a protozoan parasite that infects mammalian hosts and causes a spectrum of disease manifestations that is strongly associated with the genetic background of the host. Interleukin (IL)-6 is an acute phase proinflammatory cytokine, known in vitro to be involved in the inhibition of the generation of regulatory T cells. IL-6-deficient mice were infected with L. major, and T cell and monocyte subsets were analyzed with flow cytometry. Our data show that at the site of infection in the footpad and in the draining popliteal lymph node, numbers of regulatory T cells remain unchanged between WT and IL-6-deficient mice. However, the spleens of IL-6(-/-) mice contained fewer regulatory T cells after infection with L. major. The development of cutaneous lesions is similar between WT and IL-6-deficient mice, while parasite burden in IL-6(-/-) mice is reduced compared to WT. The development of IFN-γ or IL-10 producing T cells is similar in IL-6(-/-) mice. Despite a comparable adaptive T cell response, IL-6-deficient mice develop an earlier peak of some inflammatory cytokines than WT mice. This data indicate that the role of IL-6 in the differentiation of regulatory T cells is complex in vivo, and the effect of an absence of this cytokine can be counter-intuitive.
Assuntos
Interleucina-6/imunologia , Leishmania major/imunologia , Leishmaniose Cutânea/imunologia , Monócitos/citologia , Subpopulações de Linfócitos T/citologia , Animais , Citocinas/sangue , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Monócitos/classificação , Monócitos/imunologia , Óxido Nítrico Sintase Tipo II/biossíntese , Organismos Livres de Patógenos Específicos , Subpopulações de Linfócitos T/imunologiaRESUMO
The effective maturation of dendritic cells (DC) is complex and highly regulated and requires the presence of a variety of signals. Tumor necrosis factor (TNF) and its receptors or innate pattern recognition receptors such as the toll-like receptors have been shown to contribute to this process. DC derived from bone marrow cells in the presence of granulocyte/macrophage colony-stimulating factor can be used as a model to ascertain the contribution of different signals to DC maturation. Analysis of DC activated by addition of the mycobacterial vaccine strain Bacillus Calmette-Guérin showed that of the effector molecules studied only interleukin-10 expression was significantly reduced in TNF-negative (B6.TNF(-/-)) DC. Another effector molecule produced by DC, inducible nitric oxide synthase, was largely unchanged.
Assuntos
Vacina BCG/metabolismo , Células Dendríticas/metabolismo , Interleucina-10/biossíntese , Receptores Tipo II do Fator de Necrose Tumoral/genética , Fator de Necrose Tumoral alfa/genética , Animais , Vacina BCG/imunologia , Medula Óssea/patologia , Diferenciação Celular/genética , Células Cultivadas , Células Dendríticas/imunologia , Células Dendríticas/microbiologia , Células Dendríticas/patologia , Regulação da Expressão Gênica , Interleucina-10/genética , Interleucina-10/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase/metabolismoRESUMO
The generalized lymphoproliferative disorder (gld) mouse strain is characterized by severe splenomegaly/lymphadenopathy, the production of autoimmune antibodies, and the appearance of CD4/CD8-negative T cells. An additional TNF deficiency of gld/gld mice attenuates the course of the disorder through a yet-unknown mechanism. In this study, we could demonstrate that the reduced splenomegaly and lymphadenopathy in B6.gld/gld.TNF-/- mice were correlated with a decreased peripheral T cell proliferation rate and a delayed polyclonal activation. A comparative analysis of naïve T cells and memory/effector T cells showed an age-dependent difference in the T cell activation pattern in the spleen of B6.gld/gld and B6.gld/gld.TNF-/- mice. T cells from B6.gld/gld.TNF-/- spleens and lymph nodes showed significantly higher levels of CCR7 and CD62 ligand on their surface compared with B6.gld/gld mice when mice of the same age were compared. Additionally, we found an increased titer of the Th1 cytokine IFN-gamma in the serum of B6.gld/gld mice, whereas the concentration of IFN-gamma was markedly reduced in the serum of B6.gld/gld.TNF-/- mice. These findings support the hypothesis that increased T cell activation and proliferation in the presence of TNF contribute to the exacerbation of the gld syndrome.
Assuntos
Envelhecimento/metabolismo , Ativação Linfocitária , Transtornos Linfoproliferativos/genética , Fator de Necrose Tumoral alfa/biossíntese , Animais , Diferenciação Celular , Proliferação de Células , Receptores de Hialuronatos/metabolismo , Interferon gama/sangue , Selectina L/metabolismo , Linfonodos/imunologia , Linfonodos/patologia , Doenças Linfáticas/imunologia , Doenças Linfáticas/metabolismo , Doenças Linfáticas/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptores CCR7/metabolismo , Baço/imunologia , Baço/patologia , Esplenomegalia/imunologia , Esplenomegalia/metabolismo , Esplenomegalia/patologia , Subpopulações de Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/genéticaRESUMO
Myeloid lineage cells present in human peripheral blood include dendritic cells (DC) and monocytes. The DC are identified phenotypically as HLA-DR+ cells that lack major cell surface lineage markers for T cells (CD3), B cells (CD19, CD20), NK cells (CD56), red blood cells (CD235a), hematopoietic stem cells (CD34), and Mo that express CD14. Both DC and Mo can be phenotypically divided into subsets. DC are divided into plasmacytoid DC, which are CD11c- , CD304+ , CD85g+ , and myeloid DC that are CD11c+ . The CD11c+ DC are readily classified as CD1c+ DC and CD141+ DC. Monocytes are broadly divided into the CD14+ CD16- (classical) and CD14dim CD16+ subsets (nonclassical). A population of myeloid-derived cells that have DC characteristics, that is, HLA-DR+ and lacking lineage markers including CD14, but express CD16 are generally clustered with CD14dim CD16+ monocytes. We used high-dimensional clustering analyses of fluorescence and mass cytometry data, to delineate CD14+ monocytes, CD14dim CD16+ monocytes (CD16+ Mo), and CD14- CD16+ DC (CD16+ DC). We sought to identify the functional and kinetic relationship of CD16+ DC to CD16+ Mo. We demonstrate that differentiation of CD16+ DC and CD16+ Mo during activation with IFNγ in vitro and as a result of an allo-hematopoietic cell transplant (HCT) in vivo resulted in distinct populations. Recovery of blood CD16+ DC in both auto- and allo-(HCT) patients after myeloablative conditioning showed similar reconstitution and activation kinetics to CD16+ Mo. Finally, we show that expression of the cell surface markers CD300c, CCR5, and CLEC5a can distinguish the cell populations phenotypically paving the way for functional differentiation as new reagents become available.
Assuntos
Células Apresentadoras de Antígenos/imunologia , Biomarcadores/análise , Células Dendríticas/imunologia , Doença Enxerto-Hospedeiro/imunologia , Monócitos/imunologia , Células Mieloides/imunologia , Receptores de IgG/metabolismo , Células Apresentadoras de Antígenos/metabolismo , Antígenos de Superfície/metabolismo , Diferenciação Celular , Linhagem da Célula , Células Dendríticas/metabolismo , Proteínas Ligadas por GPI/metabolismo , Doença Enxerto-Hospedeiro/diagnóstico , Doença Enxerto-Hospedeiro/metabolismo , Antígenos HLA-DR/metabolismo , Transplante de Células-Tronco Hematopoéticas , Humanos , Lectinas Tipo C/metabolismo , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/terapia , Glicoproteínas de Membrana/metabolismo , Monócitos/metabolismo , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/terapia , Células Mieloides/metabolismo , Receptores CCR5/metabolismo , Receptores de Superfície Celular/metabolismo , Transplante HomólogoRESUMO
Monoclonal antibodies targeting co-inhibitory immune checkpoint molecules have been successful in clinical trials of both solid and hematological malignancies as acknowledged by the 2018 Nobel Prize in Medicine, however improving clinical response rates is now key to expanding their efficacy in areas of unmet medical need. Antibodies to checkpoint inhibitors target molecules on either T cells or tumor cells to stimulate T cells or remove tumor mediated immunosuppression, respectively. However, many of the well-characterized T cell immune checkpoint receptors have their ligands on antigen presenting cells or exert direct effects on those cells. Dendritic cells are the most powerful antigen presenting cells; they possess the ability to elicit antigen-specific responses and have important roles in regulation of immune tolerance. Despite their theoretical benefits in cancer immunotherapy, the translation of DC therapies into the clinic is yet to be fully realized and combining DC-based immunotherapy with immune checkpoint inhibitors is an attractive strategy. This combination takes advantage of the antigen presenting capability of DC to maximize specific immune responses to tumor antigens whilst removing tumor-associated immune inhibitory mechanisms with immune checkpoint inhibition. Here we review the expression and functional effects of immune checkpoint molecules on DC and identify rational combinations for DC vaccination to enhance antigen-specific T cell responses, cytokine production, and promotion of long-lasting immunological memory.
RESUMO
Antibody-based therapy in acute myeloid leukemia (AML) has been marred by significant hematologic toxicity due to targeting of both hematopoietic stem and progenitor cells (HSPCs). Achieving greater success with therapeutic antibodies requires careful characterization of the potential target molecules on AML. One potential target is CD300f, which is an immunoregulatory molecule expressed predominantly on myeloid lineage cells. To confirm the value of CD300f as a leukemic target, we showed that CD300f antibodies bind to AML from 85% of patient samples. While one CD300f monoclonal antibody (mAb) reportedly did not bind healthy hematopoietic stem cells, transcriptomic analysis found that CD300f transcripts are expressed by healthy HSPC. Several CD300f protein isoforms exist as a result of alternative splicing. Importantly for antibody targeting, the extracellular region of CD300f can be present with or without the exon 4-encoded sequence. This results in CD300f isoforms that are differentially bound by CD300f-specific antibodies. Furthermore, binding of one mAb, DCR-2, to CD300f exposes a structural epitope recognized by a second CD300f mAb, UP-D2. Detailed analysis of publicly available transcriptomic data indicated that CD34+ HSPC expressed fewer CD300f transcripts that lacked exon 4 compared to AML with monocytic differentiation. Analysis of a small cohort of AML cells revealed that the UP-D2 conformational binding site could be induced in cells from AML patients with monocytic differentiation but not those from other AML or HSPC. This provides the opportunity to develop an antibody-based strategy to target AMLs with monocytic differentiation but not healthy CD34+ HSPCs. This would be a major step forward in developing effective anti-AML therapeutic antibodies with reduced hematologic toxicity.
Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos Imunológicos/farmacologia , Epitopos/imunologia , Leucemia Mieloide Aguda/tratamento farmacológico , Receptores Imunológicos/imunologia , Linhagem Celular Tumoral , Humanos , Leucemia Mieloide Aguda/imunologia , Terapia de Alvo Molecular , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Receptores Imunológicos/antagonistas & inibidoresRESUMO
Acute myeloid leukemia (AML) is the most common form of adult acute leukemia with ~20,000 new cases yearly. The disease develops in people of all ages, but is more prominent in the elderly, who due to limited treatment options, have poor overall survival rates. Monoclonal antibodies (mAb) targeting specific cell surface molecules have proven to be safe and effective in different haematological malignancies. However, AML target molecules are currently limited so discovery of new targets would be highly beneficial to patients. We examined the C-type lectin receptor CD302 as a potential therapeutic target for AML due to its selective expression in myeloid immune populations. In a cohort of 33 AML patients with varied morphological and karyotypic classifications, 88% were found to express CD302 on the surface of blasts and 80% on the surface of CD34+ CD38- population enriched with leukemic stem cells. A mAb targeting human CD302 was effective in mediating antibody dependent cell cytotoxicity and was internalised, making it amenable to toxin conjugation. Targeting CD302 with antibody limited in vivo engraftment of the leukemic cell line HL-60 in NOD/SCID mice. While CD302 was expressed in a hepatic cell line, HepG2, this molecule was not detected on the surface of HepG2, nor could HepG2 be killed using a CD302 antibody-drug conjugate. Expression was however found on the surface of haematopoietic stem cells suggesting that targeting CD302 would be most effective prior to haematopoietic transplantation. These studies provide the foundation for examining CD302 as a potential therapeutic target for AML.
Assuntos
Antígenos de Neoplasias/metabolismo , Antineoplásicos Imunológicos/farmacologia , Crise Blástica , Sistemas de Liberação de Medicamentos , Lectinas Tipo C/metabolismo , Leucemia Mieloide Aguda , Células-Tronco Neoplásicas , Receptores de Superfície Celular/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Crise Blástica/tratamento farmacológico , Crise Blástica/metabolismo , Crise Blástica/patologia , Feminino , Células HL-60 , Transplante de Células-Tronco Hematopoéticas , Células Hep G2 , Humanos , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Leucemia Mieloide Aguda/terapia , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Only modest advances in AML therapy have occurred in the past decade and relapse due to residual disease remains the major challenge. The potential of the immune system to address this is evident in the success of allogeneic transplantation, however this leads to considerable morbidity. Dendritic cell (DC) vaccination can generate leukemia-specific autologous immunity with little toxicity. Promising results have been achieved with vaccines developed in vitro from purified monocytes (Mo-DC). We now demonstrate that blood DC (BDC) have superior function to Mo-DC. Whilst BDC are reduced at diagnosis in AML, they recover following chemotherapy and allogeneic transplantation, can be purified using CMRF-56 antibody technology, and can stimulate functional T cell responses. While most AML patients in remission had a relatively normal T cell landscape, those who had received fludarabine as salvage therapy have persistent T cell abnormalities including reduced number, altered subset distribution, failure to expand, and increased activation-induced cell death. Furthermore, PD-1 and TIM-3 are increased on CD4T cells in AML patients in remission and their blockade enhances the expansion of leukemia-specific T cells. This confirms the feasibility of a BDC vaccine to consolidate remission in AML and suggests it should be tested in conjunction with checkpoint blockade.
RESUMO
HLDA10 collated a panel of monoclonal antibodies (mAbs) that primarily recognised molecules on human myeloid cell and dendritic cell (DC) populations. As part of the studies, we validated a backbone of mAbs to delineate monocyte and DC populations from peripheral blood. The mAb backbone allowed identification of monocyte and DC subsets using fluorochromes that were compatible with most 'off the shelf' or routine flow cytometers. Three laboratories used this mAb backbone to assess the HLDA10 panel on blood monocytes and DCs. Each laboratory was provided with enough mAbs to perform five repeat experiments. The data were collated and analysed using Spanning-tree Progression Analysis of Density-normalised Events (SPADE). The data were interrogated for inter- and intra-laboratory variability. The results highlight the definition of DC populations using current readily available reagents. This collaborative process provides the broader scientific community with an invaluable data set that validates mAbs to leucocyte surface molecules.