Immune therapies that have used regulatory T cells (Tregs) have long been limited by difficulties in suppressing the harmful immune response in autoimmune disorders without compromising the immune system as a whole. While Tregs are the natural mechanism for maintaining immune tolerance, inducing antigen-specific Tregs safely and efficiently in patients has been difficult. Existing strategies often depend on broad immunosuppression or complex cell therapies, both of which carry risks such as infections, malignancies, or non-specific immune suppression. Now, researchers at the Nano Life Science Institute (WPI-NanoLSI) and the Faculty of Medicine at Kanazawa University have developed a potential way around these challenges using an engineered extracellular vesicle (EV) platform designed to induce antigen-specific Tregs in a controlled manner. The work, published in Drug Delivery, could be a breakthrough to creating therapies to treat autoimmune diseases that selectively target disease-related antigens while preserving protective immunity.
“A long-standing goal in immunology has been the development of therapies that suppress immune responses only toward disease-related antigens,” the researchers noted in a press release, adding that their EV-based method “may pave the way for next-generation therapies for autoimmune and allergic diseases, where unwanted immune activation must be precisely controlled.”
The concept of using EVs as delivery vehicles emerged from earlier research which demonstrated that they can carry multiple functional molecules and interact efficiently with immune cells. EVs are nanoscale particles released by cells that contain proteins, lipids, and other biological components capable of altering cell behavior. Building on their previous work that used EVs to activate antigen-specific T cells in cancer models, the team thought that a similar platform could be adapted to promote immune tolerance instead of immune activation.
To test this, the researchers engineered what they call antigen-presenting EVs, or AP-EVs. These EVs were designed to simultaneously display three signals required for Treg differentiation: peptide–major histocompatibility complex class II complexes (pMHCII) for antigen recognition, and the cytokines interleukin-2 (IL-2) and transforming growth factor-β (TGF-β). “These immunomodulatory molecules were anchored to the EV membrane via CD81 or milk fat globule-EGF factor 8 (MFG-E8) scaffolds to ensure stable and multivalent presentation,” the researchers wrote.
In the lab, the team co-cultured AP-EVs with naïve CD4⁺ T cells from antigen-specific T cell receptor–transgenic mice. The EVs induced differentiation and expansion of Foxp3⁺ Tregs, the hallmark regulatory T cell population. The induced cells expressed regulatory markers such as CD25, CTLA-4, PD-L1, and LAG-3, and they suppressed the proliferation of other T cells. The finding that the Tregs induced by AP-EVs in vitro exhibited suppressive function, pointed to the therapeutic potential of the newly developed system.
The researchers then turned to mouse models to further evaluate if this approach was effective. The data from this work showed that the AP-EVs selectively activated antigen-specific CD4⁺ T cells, but efficient induction of Foxp3⁺ Tregs in vivo required the addition of rapamycin, an mTOR inhibitor known to favor Treg differentiation. These discoveries “highlight the importance of mTOR inhibition under physiological conditions,” the researchers noted, which showed that the in vivo immune environment provides competing activation signals that can limit the induction of tolerance.
There are a number of features of EVs that distinguish them from other delivery systems previously explored to induce immune tolerance. Unlike synthetic nanoparticles or mRNA-based approaches, EVs are naturally derived and show high biocompatibility and low immunogenicity. The researchers noted that mRNA therapies can face challenges related to off-target effects and transient expression, while adoptive Treg therapies require complex ex vivo expansion and carry risks of polyclonal immunosuppression. EVs, by contrast, can present multiple functional molecules at once, a feature that “allows for the tuning of antigen specificity and immunoregulatory signals,” the team noted.
This modularity could serve drug developers well as it would allow for a cell-free treatment strategy that could be tuned according to the different needs of the autoimmune disease or allergy by simply changing the antigen loaded onto the EV. The ability to induce antigen-specific Tregs could allow for the suppression of disease-driving immune responses without compromising overall immune defense. The approach could also reduce reliance on chronic systemic immunosuppression.
Future studies to further refine this approach should focus on optimizing EV design, evaluating long-term stability and suppressive function of induced Tregs, and assessing the safety of this approach. The researchers also suggested incorporating additional inhibitory molecules into AP-EVs to mimic the effects of rapamycin, a step that could further refine antigen-specific immune tolerance therapies.
