RESUMEN
Based on the success in animal models of type 1 diabetes (T1D), clinical trials of adoptive regulatory T cell (Treg) therapy are underway using ex vivo expanded polyclonal Tregs. However, pre-clinical data also demonstrate that islet-specific Tregs are more potent than polyclonal Tregs at reversing T1D. Translation of this approach into man will require methods to generate large populations of islet-specific Tregs which, to date, has proved to be a major hurdle. Here we demonstrate the feasibility of lentiviral-mediated T cell receptor (TCR) gene transfer to confer antigen specificity on polyclonal human Tregs. Targeting has been achieved using TCRs isolated from human islet-specific and viral-specific CD4+ T cell clones. Engineered T cells demonstrated expression of ectopically-delivered TCRs, resulting in endowment of cognate antigen-specific responses. This enabled antigen-specific suppression at increased potency compared to polyclonal Tregs. However, cells transduced with islet-specific TCRs were less responsive to cognate antigen than viral-specific TCRs, and in some cases, required additional methods to isolate functional antigen-specific Tregs. This study demonstrates the potential of TCR gene transfer to develop islet-specific Treg therapies for effective treatment of T1D, but also highlights that additional optimisation may be required to achieve its full potential.
Asunto(s)
Islotes Pancreáticos/inmunología , Receptores de Antígenos de Linfocitos T/genética , Especificidad del Receptor de Antígeno de Linfocitos T/genética , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Animales , Línea Celular , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/terapia , Modelos Animales de Enfermedad , Epítopos de Linfocito T/inmunología , Orden Génico , Técnicas de Transferencia de Gen , Terapia Genética , Vectores Genéticos/genética , Humanos , Células Jurkat , Lentivirus/genética , Ratones , Transducción GenéticaRESUMEN
In the Guillain-Barré syndrome subform acute motor axonal neuropathy (AMAN), Campylobacter jejuni enteritis triggers the production of anti-ganglioside Abs (AGAbs), leading to immune-mediated injury of distal motor nerves. An important question has been whether injury to the presynaptic neuron at the neuromuscular junction is a major factor in AMAN. Although disease modeling in mice exposed to AGAbs indicates that complement-mediated necrosis occurs extensively in the presynaptic axons, evidence in humans is more limited, in comparison to the extensive injury seen at nodes of Ranvier. We considered that rapid AGAb uptake at the motor nerve terminal membrane might attenuate complement-mediated injury. We found that PC12 rat neuronal cells rapidly internalized AGAb, which were trafficked to recycling endosomes and lysosomes. Consequently, complement-mediated cytotoxicity was attenuated. Importantly, we observed the same AGAb endocytosis and protection from cytotoxicity in live mouse nerve terminals. AGAb uptake was attenuated following membrane cholesterol depletion in vitro and ex vivo, indicating that this process may be dependent upon cholesterol-enriched microdomains. In contrast, we observed minimal AGAb uptake at nodes of Ranvier, and this structure thus remained vulnerable to complement-mediated injury. These results indicate that differential endocytic processing of AGAbs by different neuronal and glial membranes might be an important modulator of site-specific injury in acute AGAb-mediated Guillain-Barré syndrome subforms and their chronic counterparts.