Multifunctional Nanoparticles Enable Efficient Oral Delivery of Biomacromolecules via Improving Payload Stability and Regulating the Transcytosis Pathway.

In oral delivery of biomacromolecules, ligand-modified nanoparticles (NPs) have emerged as a promising tool to improve the epithelial uptake of the loaded protein/peptide. Unfortunately, the stability and the transport mechanisms of the biotherapeutics during the intracellular transportation still remained unclear, leading to the poor transepithelial efficiency. Additionally, developing novel approaches to simultaneously monitor the payload bioactivity during the transport processes is veritably benefit for keeping their bioactivity. In the present study, EGP peptide (KRKKKGKGLGKKRDPCLRKYK), a ligand with high affinity to heparan sulfate proteoglycans (HSPGs), was found remarkably increasing the cellular uptake (4.5-fold) and also surprisingly achieving high transcytosis efficiency (4.2-fold) of poly(lactide- co-glycolide) NPs on Caco-2 cell monolayer. Compared with unmodified NPs (C NPs), EGP modified NPs (EGP NPs) exhibited more desirable colloidal stability within epithelia. In the subsequent study, the bioactivity of encapsulated insulin during the cellular transportation was innovatively monitored by a glucose consumption assay. Inspiringly, EGP NPs could mostly retain the bioactivity of loaded insulin whereas insulin from INS-C NPs was significantly degraded. Then the detailed mechanism study revealed that the binding of EGP to HSPGs played a vital role on NP transportation. Unlike C NPs being delivered in the endo/lysosomal pathway, EGP NPs were involved in caveolae-mediated transport, which contributes to the efficient avoidance of the lysosomal entrapment and sequentially facilitates the direct apical-to-basolateral transcytosis. The enhanced absorption of EGP NPs was confirmed in in situ intestinal loop models. Most importantly, oral administrated INS-EGP NPs generated a strong hypoglycemic response on diabetic rats with 10.2-fold and 2.6-fold increase in bioavailability compared with free insulin and INS-C NPs, respectively. The work provided an innovative strategy to monitor the payload bioactivity during the transport processes and proposed a novel aspect to increase oral bioavailability of biomacromolecules via improving payload stability and regulating the transcytosis pathway of nanocarriers.