Abstract
Dendritic cell (DC)-derived exosomes (DC EXO), natural nanoparticles of endosomal origin, are under intense scrutiny in clinical trials for various inflammatory diseases. DC EXO are eobiotic, meaning they are well-tolerated by the host; moreover, they can be custom-tailored for immune-regulatory or -stimulatory functions, thus presenting attractive opportunities for immune therapy. Previously we documented the efficacy of immunoregulatory DCs EXO (regDCs EXO) as immunotherapy for inflammatory bone disease, in an in-vivo model. We showed a key role for encapsulated TGFβ1 in promoting a bone sparing immune response. However, the on- and off-target effects of these therapeutic regDC EXO and how target signaling in acceptor cells is activated is unclear. In the present report, therapeutic regDC EXO were analyzed by high throughput proteomics, with non-therapeutic EXO from immature DCs and mature DCs as controls, to identify shared and distinct proteins and potential off-target proteins, as corroborated by immunoblot. The predominant expression in regDC EXO of immunoregulatory proteins as well as proteins involved in trafficking from the circulation to peripheral tissues, cell surface binding, and transmigration, prompted us to investigate how these DC EXO are biodistributed to major organs after intravenous injection. Live animal imaging showed preferential accumulation of regDCs EXO in the lungs, followed by spleen and liver tissue. In addition, TGFβ1 in regDCs EXO sustained downstream signaling in acceptor DCs. Blocking experiments suggested that sustaining TGFβ1 signaling require initial interaction of regDCs EXO with TGFβ1R followed by internalization of regDCs EXO with TGFβ1-TGFβ1R complex. Finally, these regDCs EXO that contain immunoregulatory cargo and showed biodistribution to lungs could downregulate the main severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) target receptor, ACE2 on recipient lung parenchymal cells via TGFβ1 in-vitro. In conclusion, these results in mice may have important immunotherapeutic implications for lung inflammatory disorders.
Highlights
Exosomes (EXO) are nanoparticles of endosomal origin secreted by all cells, including dendritic cells (DCs), the most potent antigen presenting cells and “directors” of immune response [1]
DC EXO subtypes are complex nano-particles, with on-target functions typically consistent with the phenotype of source DCs. This was previously reported by our group and includes regulation by regDC EXO of inflammatory cytokines, and the reprogramming of DC maturation and Treg-Th17 cell effector differentiation [5]
We focused on in-depth proteomic LC-MS/MS analysis, to identify both on- and offtarget proteins that could lead to unintended consequences of DC EXO therapy
Summary
Exosomes (EXO) are nanoparticles of endosomal origin secreted by all cells, including dendritic cells (DCs), the most potent antigen presenting cells and “directors” of immune response [1]. EXO contain proteins, nucleic acids and lipid cargo that mediate intercellular communication and signaling They are secreted into tissues and body fluids and can act locally or from at a distance [2]. We have previously reported important aspects of the immunobiology and functions of DC-derived EXO subtypes, isolated from DCs at distinct stages of maturation. Most notably among these subtypes are immune-regulatory (regDCs EXO), loaded with TGFb1 and IL10 and deficient in costimulatory molecules. Immature or immune-null DC exo, (iDCs EXO) and EXO from matured DCs, called immune-stimulatory (stimDCs EXO), were characterized for immune functions [5] The stability of these EXO, their ability to protect their cargo, and be retained at inflamed mucosal sites and inhibit inflammatory bone loss has been documented. How regDCs EXO modulate cytokine signaling in recipient cells [5], needs further investigations
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