RESUMEN
Monoclonal antibodies are among the most promising therapeutic agents for treating cancer. Therapeutic cancer antibodies bind to tumor cells, turning them into targets for immune-mediated destruction. We show here that this antibody-mediated killing of tumor cells is limited by a mechanism involving the interaction between tumor cell-expressed CD47 and the inhibitory receptor signal regulatory protein-α (SIRPα) on myeloid cells. Mice that lack the SIRPα cytoplasmic tail, and hence its inhibitory signaling, display increased antibody-mediated elimination of melanoma cells in vivo. Moreover, interference with CD47-SIRPα interactions by CD47 knockdown or by antagonistic antibodies against CD47 or SIRPα significantly enhances the in vitro killing of trastuzumab-opsonized Her2/Neu-positive breast cancer cells by phagocytes. Finally, the response to trastuzumab therapy in breast cancer patients appears correlated to cancer cell CD47 expression. These findings demonstrate that CD47-SIRPα interactions participate in a homeostatic mechanism that restricts antibody-mediated killing of tumor cells. This provides a rational basis for targeting CD47-SIRPα interactions, using for instance the antagonistic antibodies against human SIRPα described herein, to potentiate the clinical effects of cancer therapeutic antibodies.
Asunto(s)
Citotoxicidad Celular Dependiente de Anticuerpos , Antígeno CD47/metabolismo , Neoplasias Mamarias Experimentales/patología , Receptores Inmunológicos/metabolismo , Animales , Anticuerpos Monoclonales Humanizados/farmacología , Citotoxicidad Celular Dependiente de Anticuerpos/efectos de los fármacos , Antineoplásicos/farmacología , Antígeno CD47/inmunología , Femenino , Citometría de Flujo , Neoplasias Mamarias Experimentales/inmunología , Neoplasias Mamarias Experimentales/metabolismo , Ratones , Ratones Endogámicos C57BL , Receptores Inmunológicos/inmunología , Transducción de Señal , TrastuzumabRESUMEN
Neutrophils potently kill tumour cells in the presence of anti-tumour antibodies in vitro. However, for in vivo targeting, the neutrophils need to extravasate from the circulation by passing through endothelial barriers. To study neutrophil migration in the presence of endothelial cells in vitro, we established a three-dimensional collagen culture in which SK-BR-3 tumour colonies were grown in the presence or absence of an endothelial barrier. We demonstrated that - in contrast to targeting FcγR on neutrophils with mAbs - targeting the immunoglobulin A Fc receptor (FcαRI) instead triggered neutrophil migration and degranulation leading to tumour destruction, which coincided with release of the pro-inflammatory cytokines interleukin (IL)-1ß and tumour necrosis factor (TNF)-α. Interestingly, neutrophil migration was enhanced in the presence of endothelial cells, which coincided with production of significant levels of the neutrophil chemokine IL-8. This supports the idea that stimulation of neutrophil FcαRI, but not FcγR, initiates cross-talk between neutrophils and endothelial cells, leading to enhanced neutrophil migration towards tumour colonies and subsequent tumour killing.
Asunto(s)
Antígenos CD/inmunología , Neoplasias de la Mama/inmunología , Quimiotaxis de Leucocito/inmunología , Células Endoteliales de la Vena Umbilical Humana/inmunología , Neutrófilos/inmunología , Receptores Fc/inmunología , Neoplasias de la Mama/patología , Comunicación Celular/inmunología , Línea Celular Tumoral , Células Endoteliales de la Vena Umbilical Humana/citología , Humanos , Inmunidad Innata/inmunología , Inmunohistoquímica , Interleucina-1beta/análisis , Interleucina-1beta/inmunología , Neutrófilos/citología , Factor de Necrosis Tumoral alfa/análisis , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
To elucidate the mechanisms of glomerulonephritis, including Goodpasture's syndrome, mouse models are used that use heterologous Abs against the glomerular basement membrane (GBM) with or without preimmunization with foreign IgG from the same species. These studies have revealed the requirement of either FcgammaR or complement, depending on the experimental model used. In this study, we provide evidence that both FcgammaR and complement are obligatory for a full-blown inflammation in a novel attenuated passive model of anti-GBM disease. We demonstrate that administration of subnephritogenic doses of rabbit anti-GBM Abs followed by a fixed dose of mouse mAbs to rabbit IgG, allowing timing and dosing for the induction of glomerulonephritis, resulted in reproducible complement activation via the classical pathway of complement and albuminuria in wild-type mice. Because albuminuria was absent in FcR-gamma-chain(-/-) mice and reduced in C3(-/-) mice, a role for both FcgammaR and complement is postulated. Because C1q(-/-) and C4(-/-) mice lacking a functional classical and lectin pathway did develop albuminuria, we suggest involvement of the alternative pathway of complement. Anti-GBM glomerulonephritis occurs acutely following the administration of mouse anti-rabbit IgG, and proceeds in a chronic fashion dependent on both FcgammaR and complement. This novel attenuated model allows elucidating the relative contribution of different mediator systems of the immune system to the development of renal injury, and also provides a platform for the assessment of different treatment protocols and evaluation of drugs that ultimately may be beneficial for the treatment of anti-GBM mediated glomerulonephritides.
Asunto(s)
Enfermedad por Anticuerpos Antimembrana Basal Glomerular/inmunología , Nefritis/inmunología , Receptores de Complemento/fisiología , Receptores de IgG/fisiología , Albuminuria/etiología , Animales , Enfermedad por Anticuerpos Antimembrana Basal Glomerular/etiología , Enfermedad por Anticuerpos Antimembrana Basal Glomerular/patología , Autoanticuerpos , Activación de Complemento , Inmunoglobulina G/administración & dosificación , Inflamación , Ratones , Nefritis/patologíaRESUMEN
BACKGROUND & AIMS: Development of liver metastases is a frequent complication in patients with colorectal cancer (CRC), even after successful resection of the primary tumor. As such, post-operative adjuvant therapies that aim to eliminate residual disease after surgery may improve patient outcome. METHODS: We used a colon carcinoma liver metastases model, in which CC531s colon carcinoma cells are injected into the portal circulation by a surgical procedure. As injected tumor cells are arrested in the liver, this model is suitable for investigating the interaction of tumor cells with the liver microenvironment. By administering tumor specific monoclonal antibodies (mAb) directly post-operatively, we were able to determine the effect of antibody therapy on eradication of arrested tumor cells and subsequent liver metastases outgrowth. RESULTS: We showed that post-operative treatment with tumor specific monoclonal antibodies (mAb) prevents liver metastases outgrowth. Antibody-dependent phagocytosis (ADPh) was the main mechanism involved, as enhanced uptake of tumor cells by innate mononuclear phagocytes in the liver was observed after mAb therapy. Furthermore, Kupffer cells (KC) were identified as the most prominent effector cells, as depletion of KC abolished therapeutic efficacy. This was partly compensated by monocytes when animals were treated with a high mAb dose, but monocytes were unable to phagocytose tumor cells when rats were treated with low mAb doses. CONCLUSIONS: The finding that KC and monocytes can eliminate tumor cells through ADPh has important and promising clinical implications for designing new adjuvant therapies for patients undergoing CRC resection.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Hepáticas/tratamiento farmacológico , Animales , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/cirugía , Terapia Combinada , Humanos , Leucocitos Mononucleares , Neoplasias Hepáticas/prevención & control , Neoplasias Hepáticas/secundario , Neoplasias Hepáticas/cirugía , Neoplasias Hepáticas Experimentales , Ratones , Fagocitos , Fagocitosis , Resultado del TratamientoRESUMEN
BACKGROUND: Anti-glomerular basement membrane (anti-GBM) disease often results in end-stage renal failure despite therapy with plasma exchange and immunosuppressive drugs. The newly discovered streptococcal enzymes IgG-degrading enzyme of S.pyogenes (IdeS) and endoglycosidase S (EndoS) act with remarkable specificity on circulating IgG. In this study, we investigate their ability in vivo to prevent damage mediated by kidney-bound antibodies in a mouse model of anti-GBM disease. METHODS: Anti-GBM disease was induced in mice by injection of subnephritogenic doses of rabbit anti-mouse GBM, followed a week later by injection of monoclonal mouse anti-rabbit IgG antibodies. By administrating IdeS or EndoS as fusion partners with GST between these antibody injections, we tested their ability to prevent damage by acting on kidney-bound rabbit anti-GBM. Control animals received placebo injections. RESULTS: All animals in the positive control groups developed severe albuminuria immediately after the second antibody injection (mean, 2.51 mg/24 h; range, 0.13-8.20). This was significantly diminished by EndoS (1.3 +/- 1.3 mg/24 h) and completely prevented by IdeS (0.017 +/- 0.014 mg/24 h). Immunofluorescence studies showed that IdeS treatment effectively removed the Fc fragments of the rabbit IgG. This was accompanied by a significant reduction of the deposition of the complement components C3 and C1q, and this diminished the recruitment of leukocytes to the glomeruli. CONCLUSION: IdeS degrades IgG bound to the GBM in vivo, thereby preventing renal damage in this animal model. Most likely, IdeS would degrade both circulating and kidney-bound anti-GBM in patients with Goodpasture's disease. Whether this would lead to a halt in disease progression and a better prognosis remains to be determined.
Asunto(s)
Enfermedad por Anticuerpos Antimembrana Basal Glomerular/tratamiento farmacológico , Proteínas Bacterianas/uso terapéutico , Glicósido Hidrolasas/uso terapéutico , Animales , Enfermedad por Anticuerpos Antimembrana Basal Glomerular/inducido químicamente , Enfermedad por Anticuerpos Antimembrana Basal Glomerular/patología , Autoanticuerpos/efectos adversos , Modelos Animales de Enfermedad , Inmunoglobulina G/sangre , Glomérulos Renales/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Resultado del TratamientoRESUMEN
Many patients with colorectal cancer will develop liver metastases, even after successful surgical removal of the primary tumor at a time at which no visible metastases are present. We previously demonstrated that surgery--although mandatory--paradoxically enhances the risk of developing liver metastases. Because Ab therapy has been acknowledged as a successful strategy to treat malignancies, we studied the potential of postoperative adjuvant Ab therapy to prevent outgrowth of liver metastases. Treatment with murine anti-gp75 (TA99) mAb completely prevented outgrowth of B16F10 liver metastases in over 90% of mice. Therapeutic efficacy was maintained in either C1q- or complement receptor 3-deficient mice but was completely abrogated in FcR gamma-chain knockout mice. This indicates that the classical complement pathway was not essential, but interaction with activatory Fc gammaR was necessary for successful therapy. TA99-treatment was still effective in Fc gammaRI(-/-), Fc gammaRIII(-/-), Fc gammaRI/III(-/-), and Fc gammaRI/II/III(-/-) mice, suggesting an important role for Fc gammaRIV. However, wild-type mice that were treated with TA99 Abs and an Fc gammaRIV blocking Ab (mAb 9E9) were protected against development of liver metastases as well. Only when both Fc gammaRI and Fc gammaRIV functions were simultaneously inhibited, TA99-mediated curative Ab treatment was abrogated, indicating functional redundancy between both IgG receptors in the liver. Furthermore, depletion of liver macrophages (Kupffer cells) reduced the efficacy of Ab therapy, supporting that Kupffer cells are involved as effector cells. Importantly, since Ab treatment almost completely prevented development of liver metastases, postoperative adjuvant Ab therapy may help to improve patient prognosis.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Neoplasias Hepáticas/prevención & control , Neoplasias Hepáticas/secundario , Receptores de IgG/inmunología , Animales , Neoplasias Colorrectales/patología , Citometría de Flujo , Macrófagos del Hígado/inmunología , Macrófagos del Hígado/metabolismo , Ratones , Ratones Noqueados , Receptores de IgG/metabolismoRESUMEN
Dendritic cells (DC) are professional antigen presenting cells that can induce and regulate adaptive immune responses. For that reason, DC are attractive candidates for vaccination strategies. Recently, expression of the IgA Fc receptor (FcalphaRI, CD89) was observed on DC, which activation led to DC maturation. We have investigated the potential of DC FcalphaRI as a target molecule for vaccination against cancer. FcalphaRI expression was observed on human blood myeloid DC. Furthermore, expression of FcalphaRI was low on immature DC, cultured from either human monocytes or FcalphaRI transgenic (Tg) mouse bone marrow cells. Addition of TNF-alpha to culture regimes of both human and mouse DC led to more semi-mature DC, on which FcalphaRI expression was slightly upregulated. FcalphaRI on both human and FcalphaRI Tg mouse DC was internalized after receptor crosslinking. Antigen presentation, measured in FcalphaRI Tg mouse DC, was however minimal. As antigen presentation is crucial to elicit effective T cell responses, these data suggest that targeting of DC FcalphaRI is not optimal for DC vaccination strategies.
Asunto(s)
Presentación de Antígeno/inmunología , Antígenos CD/fisiología , Células Dendríticas/inmunología , Inmunoglobulina A/metabolismo , Receptores Fc/fisiología , Animales , Antígenos CD/genética , Línea Celular , Humanos , Ratones , Ratones Transgénicos , Receptores Fc/genéticaRESUMEN
Traditionally IgA has been regarded as a non-inflammatory antibody, which inhibits adhesion of micro-organisms to the mucosal wall without initiation of inflammatory responses. Recently, however, a dichotomy has been suggested between the actions of secretory IgA (SIgA), which is present at mucosal sites, and serum IgA. SIgA exerts its function as first line of defence by limiting invasion of pathogens. Serum IgA in turn may be engaged in inflammatory responses after breaching of mucosal wall integrity. Several receptors for IgA have been described. However, the-as yet-best characterized prototypic Fc receptor for IgA, FcalphaRI (CD89), is the most likely candidate for initiation of inflammatory responses, as it binds poorly to SIgA, but vigorously triggers potent effector functions upon binding to serum IgA. Here, new insights in IgA-FcalphaRI binding are described and the functional implications of these interactions are discussed.
Asunto(s)
Antígenos CD/inmunología , Inmunidad Mucosa/inmunología , Inmunoglobulina A/inmunología , Receptores Fc/inmunología , Animales , Antígenos CD/fisiología , Enfermedades Transmisibles/inmunología , Humanos , Inmunidad Mucosa/fisiología , Receptores Fc/fisiologíaRESUMEN
Neutrophil-mediated tumor cell lysis is more efficiently triggered by FcalphaRI (CD89), than by FcgammaRI (CD64). This difference is most evident in immature neutrophils in which FcgammaRI-mediated tumor cell lysis is absent. In this study, we show that FcR gamma-chain-dependent functions (such as Ab-dependent cellular cytotoxicity and respiratory burst), as well as signaling (calcium mobilization and MAPK phosphorylation), were potently triggered via FcalphaRI, but not via FcgammaRI, in immature neutrophils. Internalization, an FcR gamma-chain-independent function, was, however, effectively initiated via both receptors. These data suggest an impaired functional association between FcgammaRI and the FcR gamma-chain, which prompted us to perform coimmunoprecipitation experiments. As a weaker association was observed between FcgammaRI and FcR gamma-chain, compared with FcalphaRI and FcR gamma-chain, our data support that differences between FcalphaRI- and FcgammaRI-mediated functions are attributable to dissimilarities in association with the FcR gamma-chain.
Asunto(s)
Antígenos CD/metabolismo , Neutrófilos/inmunología , Receptores Fc/metabolismo , Receptores de IgG/metabolismo , Animales , Médula Ósea/inmunología , Línea Celular Tumoral , Humanos , Inmunoprecipitación , Ratones , Estallido Respiratorio , Transducción de SeñalRESUMEN
Antitumor Abs are promising therapeutics for cancer. Currently, most Ab-based therapies focus on IgG Ab, which interact with IgG FcR (FcgammaR) on effector cells. In this study, we examined human and mouse neutrophil-mediated tumor cell lysis via targeting the IgA FcR, FcalphaRI (CD89), in more detail. FcalphaRI was the most effective FcR in triggering tumor cell killing, and initiated enhanced migration of neutrophils into tumor colonies. Importantly, immature neutrophils that are mobilized from the bone marrow upon G-CSF treatment efficiently triggered tumor cell lysis via FcalphaRI, but proved incapable of initiating tumor cell killing via FcgammaR. This may provide a rationale for the disappointing results observed in some earlier clinical trials in which patients were treated with G-CSF and antitumor Ab-targeting FcgammaR.