Chlorzoxazone, a small molecule drug, augments immunosuppressive capacity of mesenchymal stem cells via modulation of FOXO3 phosphorylation

Nowadays, immune diseases are a large burden in healthcare. Mesenchymal stem cells (MSCs) have prominent ability in immunomodulation and have been applicated on treating many immune-related diseases. However, the clinical outcomes can be disparate and sometimes completely counterproductive beyond explanation of cell heterogeneity. The theory of immunomodulation plasticity in MSCs has then emerged to explain that MSCs can be induced into proinflammatory MSC1 or anti-inflammatory MSC2 responding to different immune environment. It would be safer and more efficient if we could induce MSCs into a certain immune phenotype, in most cases MSC2, prior to medical treatment. In this study, we screened and identified a classical FDA-approved drug, chlorzoxazone (CZ). Unlike traditional method induced by IFN-γ, CZ can induce MSC into MSC2 phenotype and enhance the immunosuppressive capacity without elevation of immunogenicity of MSCs. CZ-treated MSCs can better inhibit T cells activation and proliferation, promote expression of IDO and other immune mediators in vitro, and alleviate inflammatory infiltration and tissue damage in acute kidney injury rat model more effectively. Moreover, we discovered that CZ modulates phosphorylation of transcriptional factor forkhead box O3 (FOXO3) independent of classical AKT or ERK signaling pathways, to promote expression of downstream immune-related genes, therefore contributing to augmentation of MSCs immunosuppressive capacity. Our study established a novel and effective approach to induce MSC2, which is ready for clinical application.