In vivofate of systemically administered encapsulin protein nanocages and implications for their use in targeted drug delivery

Abstract

AbstractEncapsulins, self-assembling protein nanocages derived from prokaryotes, are promising nanoparticle-based drug delivery systems (NDDS). However, thein vivobehavior and fate of encapsulins are poorly understood. In this pre-clinical study, we probe the interactions between the model encapsulin fromThermotoga maritima(TmEnc) and key biological barriers encountered by NDDS. Here, a purified TmEnc formulation that exhibited colloidal stability, storability, and blood compatibility was intravenously injected into BALB/c mice. TmEnc had an excellent nanosafety profile, with no abnormal weight loss or gross pathology observed, and only temporary alterations in toxicity biomarkers detected. Notably, TmEnc demonstrated immunogenic properties, inducing the generation of nanocage-specific IgM and IgG antibodies, but without any prolonged pro-inflammatory effects. An absence of antibody cross-reactivity also suggested immune-orthogonality among encapsulins systems. Moreover, TmEnc formed a serum-derived protein corona on its surface which changed dynamically and appeared to play a role in immune recognition. TmEnc’s biodistribution profile further revealed its sequestration from the blood circulation by the liver and then biodegraded within Kupffer cells, thus indicating clearance via the mononuclear phagocyte system. Collectively, these findings provide critical insights into how encapsulins behavein vivo,thereby informing their future design, modification, and application in targeted drug delivery.