Phosphatidylserine-rich liposomes to tackle autoimmunity. En route to translationality

Resumen

Autoimmune diseases are caused by defective immunological tolerance, and reportedly affect up to 10% of the global population. In the last years, current medical interventions have transformed these disorders into chronic and manageable, but they still entail high rates of morbidity and mortality. Hence, there is an urgent need to develop therapies capable of restoring the breach of tolerance selectively, which halt the autoimmune aggression and allow the regeneration of the targeted tissue. In physiological conditions, the phagocytosis of apoptotic cells performed by phagocytes such as dendritic cells (DCs) —a process termed efferocytosis— prompts the acquisition of tolerogenic features and the ability to restore tolerance. Indeed, a cell immunotherapy consisting of DCs rendered tolerogenic (tolDCs) by apoptotic β-cell efferocytosis arrested the autoimmune attack against β-cells in an experimental model of type 1 diabetes (T1D). However, in light of the hurdles in obtaining and standardising human autologous apoptotic β-cells for its implementation in the clinics, a nanotherapeutic strategy based on liposomes mimicking apoptotic cells was designed. The fundamental characteristics of these synthetic vesicles are: a high percentage of phosphatidylserine (PS) —phospholipid unique to the apoptotic cell membrane—, diameter superior to 500 nm, negative charge and efficient encapsulation of insulin peptides. Importantly, this strategy was equally effective in inducing tolDCs and blunting β-cell autoimmunity as the immunotherapy based on apoptotic cells. The hypothesis of this work is that autoantigen-loaded PS-liposomes can re-establish tolerance in several antigen-specific autoimmune diseases through the induction of tolDCs and the expansion of regulatory T lymphocytes, and that they have translational potential to tackle human autoimmune disorders. The main aim of the present work has been to characterise the tolerogenic potential of PS-liposomes globally. To this end, different autoantigenic peptides relevant in autoimmune diseases have been efficiently encapsulated into PS-liposomes, without difficulties in preserving their appropriate diameter and charge, thus demonstrating the versatility of the therapy to different autoimmune pathologies. In the experimental model of T1D, the administration of PS-liposomes causes the expansion of clonal CD4+ regulatory T cells and CD8+ T cells, which contribute to the long-term re-establishment of tolerance. Moreover, in the same model, the biocompatibility and safety of the final product have been confirmed given its optimal tolerability. Furthermore, PS-liposomes have been adapted to the experimental multiple sclerosis model by merely replacing the encapsulated autoantigen. In this model, PS-liposomes elicit the generation of tolDCs and decrease the incidence and severity of the disease correlating with an increase in the frequency of regulatory T cells, a fact that validates the potential of PS-liposomes to serve as a platform for tolerance re-establishment in different autoimmune diseases. Finally, considering its future clinical implementation, the effect of the PS-liposomes therapy has been determined in human DCs obtained from patients with T1D. In DCs from adult patients, PS-liposomes are efficiently phagocyted by DCs with rapid kinetics dependent on the presence of PS, and this induces a tolerogenic transcriptome, phenotype and functionality that are similar to those observed in experimental models. However, DCs from paediatric patients display defects in their phagocytic capacity correlating with the time of disease progression, albeit their phenotype and immunoregulatory gene expression after PS-liposomes phagocytosis point to an optimal tolerogenic ability. In conclusion, the liposomal immunotherapy herein described, which is based on efferocytosis as a powerful tolerance-inducing mechanism, achieves apoptotic mimicry in a simple, safe and efficient manner. Additionally, liposomes offer advantages in terms of production and standardisation. Therefore, PS-liposomes possess translational potential and constitute an encouraging strategy to restore immunological tolerance in antigen-specific autoimmune diseases.