Design Of Biomimetic Rod-shaped Vehicles With Dual PEG Conformation And Research On In Vivo Delivery Mechanisms

Oral administration of pharmaceuticals is a more ideal,acceptable route that possesses various advantages including user friendly,convenience of dose control,good patient complicane,and economical.For most pharmaceuticals that have poor stability,low solubility,and limited permeability,however,it is hard for them to be absorbed by intestines due to the grastraintestinal environment.Although micro-and nano-sized drug vehicles bring promising future for oral delivery of these drugs,the complex physiological barriers in intestinal mucosa severely hamper the transportation and delivery effieiency of drug vehicles.The mucus layer and the epithelia in intestinal mucosa form the major biological barriers.Mucin is the main composition of mucus,which presents to be a cross-linked,fibrous system,forming a biological hydrogel-like mesh barrier.The pore sizes are largely ranging from 100 nm to 300 nm,which can severely prevent the penetration of nanoparticles.Moreover,mucin itself has strong adhesive affinities that can trap foreign particles and transfer them to the lower intestine along with the intestinal peristalsis and renewal of mucus,leaving the particles away from main absorption sites.Intestinal epithelia is a rigorous barrier covered by mucus,which is impermeable to vehicles with sizes ranging from tens to hundreds of nanometers,requiring vehicles to be taken up by cells before transcellular transport.More challengingly,efficient mucus penetration and cellular uptake have contradictory preferences in terms of the characteristics of vehicles.Therefore,it is necessary to develop more rational approaches to balancing the delivery efficiency of vehicles when facing multiple barriers.Many biological effects and natural phenomena can provide inspirations of developing drug delivery systems.Researches have developed various bio-mimetic vehicles based on enlightenment given by structures and functions of eukaryotic cells,erythrocytes,and viruses.We noticed that there exists numerous microbiomes in the human gastrointestinal tract,which are mostly in rod or rod-like shape.In addition,some pathogens such as Helicobactor pylori have flagella and fimbriae,which are polymeric proteins with different conformation,on their surface.Long Flagella can help pathogens to efficiently penetrate mucus layer,while short fimbriae can facilitate the adhesion of germs to cell membrane and realize invasion.These biological phenomena give us enlightenment that it may be possible to develop a rod-shaped vehicle with controlled polymeric conformation on its surface,consequently realizing efficient overcoming of intestinal mucosal barriers.In this project,we first designed rod-shaped vehicle that mimicked the shape of gut microbiomes to improve the mucus-penetrating ability.Via a liquid crystal-template mechanism,we fabricated mesoporous silica nanospheres(NS)and nanorods(NR)with different aspect ratios(ARs).They had similar diameters and surface charges.Superior diffusivity of rod-shaped vehicles in mucus over spheres was verified to be possible by multiple particle tracking,and there existed an optimal AR(about 3)that had the highest diffusivity.Moreover,in the comparative studies,we discovered that no matter in hydroxyethyl cellulose,which has mesh-like structure but is nonadhesive,or in glycerin,which has a high viscosity but no network,the diffusivities of vehicles monotonically decreased as the AR increased.We concluded that the special characteristics of mucus made it a medium that violate the conventional diffusing law,giving rod-shaped vehicles the opportunitiy to display their advantages.After optimization of the rod vehicle with a proper AR,we then decorated polymers on it.Polyethylene glycol(PEG)is a sort of polymer that has been extensively adopted for surface modification of drug molecules and vehicles in clinical.Recently,it has been revealed that with the dense PEG coating on particle surfaces,the molecular conformation of PEG can transform from packed regime to extended regime because of intermolecular interactions.The extended regime can greatly reduce the adhesive interactions with mucin and hence improve the mucus penetration of vehicles.However,this high PEG density has been criticized for its severe inhibition on cellular uptake,which contradicts its functions in mucus penetration.Considering the above-mentioned facts,we purposed a germ-mimetic structure that had PEG molecules with both conformation on a single particle,which might solve the controversial issues concerning the intestinal multiple barriers and materially improve the delivery efficiency of vehicles.By referring to the synthetic mechanisms and morphogical observations of NR,we noticed that the porous structures specifically distributed on the tips of NR,rather than the uniform distribution around NS.It was further discovered that the hydroxyl distribution on rod tips was significantly higher than that on rod bodies detected by atomic force microscope(AFM)with chemical modification.Since hydroxyls are the major groups for chemical reaction on the surface of silica,the anisotropic distribution of hydroxyls could lead to anisotropic PEG densities during PEGylation and consequently caused different molecular conformation.After modifying the vehicles with various PEG degrees,AFM test was performed under liquid condition.It was uncovered that with a 4% PEG degree,NR exhibited extended PEG conformation at tips while packed regime around the body.All of the NR with other PEG degrees and NS showed mono molecular conoformation.Therefore,we successfully constructed germ-mimetc nanoparticles(GMNP)with dual conformational PEG.In vitro studies showed that GMNP had a strong diffusing ability and could maintain a high efficiency in cellular uptake.In the mucus-secreting E12 cell model of multiple barriers,GMNP displayed the best transport rate.Mechanism studies revealed that GMNP moved in the mucus in a “hopping” pattern while firmly adhered to the cell membrane via its body during the uptake.Notably,the rod body of GMNP had an adhesive force with cell membrane about 35.7-fold higher than that of the rod tip.In vivo oral delivery studies further showed that GMNP could rapidly penetrate the mucus layer,which led to a good intestinal retention,and could be largely absorbed by intestinal villi.Using saquinavir(SQV),a kind of BCS IV pharmaceutical,as the model drug,we showed that GMNP imporved its oral bioavailability by 21.9 fold.Additionally,we also showed that this biomimetic design could also work for tumor-targeting delivery where similar multiple biological barriers exist.Meanwhile,there are differences in the circulation time and biodistribution between GMNP and other isotropic vehicles,which may deserve future investigations.Via mimicking the rod-like shape and surface molecular structure of germs,the developed drug vehicles in this work efficiently overcame the mucus and epithelial barriers in intestinal mucosa,greatly improving the oral bioavailability of drugs.This work may offer new ideas and guidelines to the future development of oral delivery systems.It also bring enlightenments to the design of vehicles regarding other delivery routes and modification of other polymers.