Serum albumin-fused interleukin-10 prevents neuroinflammation by promoting immunoregulation in the secondary lymphoid organs and limiting immune cell infiltration in the spinal cord

Abstract Interleukin-10 (IL-10) is a potent immunoregulatory cytokine that suppresses pro-inflammatory cytokine production, reduces antigen presentation by myeloid cells, promotes M2 macrophage polarization, and inhibits T cell activation. Despite these well-established immunoregulatory functions, efforts to harness recombinant IL-10 therapeutically have been limited by its short plasma half-life and poor retention in the secondary lymphoid organs (SLOs), key sites of autoreactive T cell priming in autoimmune disease. Previously, we engineered a fusion of serum albumin and IL-10 (SA-IL-10) with extended half-life and enhanced exposure in the SLOs following intravenous administration. Here, we integrate human transcriptomic analyses and a murine model of neuroinflammation, experimental autoimmune encephalomyelitis (EAE), to investigate how sustained IL-10 exposure in the SLOs modulates immune responses under inflammatory conditions. Human single-cell RNA sequencing analyses revealed reduced IL-10 expression alongside increased IL-10 receptor expression across multiple immune cell populations in treatment-naïve patients with multiple sclerosis (MS), motivating the investigation of IL-10–based immunomodulatory strategies. Prophylactic SA-IL-10 administration prevented the development and progression of EAE with superior efficacy to wild type IL-10 and comparable protection to fingolimod, an FDA-approved MS therapy. Immunophenotyping of the SLOs revealed that SA-IL-10 suppressed pathogenic, antigen-specific RORγt + Foxp3 - T H 17 T cells, CD86 + M1-like macrophages, CD86 + dendritic cells, and pro-inflammatory cytokine production, while expanding immunoregulatory CD206 + M2-like macrophages and increasing the frequency of multiple checkpoint markers (CTLA-4, PD-1, TIGIT, ICOS) on GATA3 + Foxp3 - T H 2 cells. Despite the absence of direct central nervous system targeting, SA-IL-10 treatment also reduced the infiltration of macrophages, dendritic cells, and CD4 + T cells into the spinal cord. Repeated SA-IL-10 administration was well tolerated, as treated EAE mice gained significantly more body weight over the course of treatment compared to PBS- and WT IL-10-treated controls, and exhibited plasma biochemistry parameters comparable to control animals at study endpoint. Together, these findings demonstrate that increasing IL-10 exposure in the SLOs suppresses neuroinflammation by promoting immunoregulation. One Sentence Summary Subcutaneously administered serum albumin-fused interleukin-10 prevents experimental autoimmune encephalomyelitis by suppressing pathogenic T H 17 cells and pro-inflammatory myeloid cells in the secondary lymphoid organs and spinal cord, while expanding immunoregulatory cells in the secondary lymphoid organs.