A fusion protein that targets antigen-loaded extracellular vesicles to B cells enhances antigen-specific T cell expansion

Extracellular vesicles (EVs) have the potential to modulate immune responses via their cargo molecules and are being explored as vehicles in cancer immunotherapy. Dendritic cell-derived EVs can induce antigen-specific immune responses leading to reduced tumor burden. This response was shown to depend partially on B cells. EVs can be targeted to certain cells or tissues, and EVs from Epstein-Barr Virus (EBV) infected cells were shown to carry the EBV glycoprotein GP350 on their surface and target human CD21 (hCD21) on B cells. We therefore investigated whether targeting EVs to B cells via this mechanism could improve antigen-specific immune responses. A soluble fusion protein containing the phosphatidylserine-binding domain (C1C2) of lactadherin and hCD21-binding domain (D123) of GP350 was used to decorate and target EVs to B cells. D123-decorated EVs increased in vitro B cell targeting 5-fold compared to EVs decorated with a non-targeting control protein or undecorated EVs. Furthermore, in vivo, D123-decoration did not alter the biodistribution of EVs across organs but specifically targeted them to B cells in the spleen, blood and lymph nodes of hCD21-transgenic mice. Immunization with hCD21-targeted, OVA-loaded EVs resulted in a higher percentage of antigen-specific CD8⁺ T cells compared to untargeted EVs. Our data show that D123-decorated EVs efficiently target B cells and improve antigen-specific T cell responses in vivo, which could be explored in future therapeutic applications.