Jin-Wook Yoo

Bio-inspired, bioengineered and biomimetic drug delivery carriers.

Synthetic carriers such as polymer and lipid particles often struggle to meet clinical expectations. Natural particulates — that range from pathogens to mammalian cells — are therefore worth examining in more depth, as they are highly optimized for their specific functions in vivo and possess features that are often desired in drug delivery carriers. With a better understanding of these biological systems, in conjunction with the availability of advanced biotechnology tools that are useful for re-engineering the various natural systems, researchers have started to exploit natural particulates for multiple applications in the delivery of proteins, small interfering RNA and other therapeutic agents. Here, we review the natural drug delivery carriers that have provided the basis and inspiration for new drug delivery systems.

Particle shape enhances specificity of antibody-displaying nanoparticles

Monoclonal antibodies are used in numerous therapeutic and diagnostic applications; however, their efficacy is contingent on specificity and avidity. Here, we show that presentation of antibodies on the surface of nonspherical particles enhances antibody specificity as well as avidity toward their targets. Using spherical, rod-, and disk-shaped polystyrene nano- and microparticles and trastuzumab as the targeting antibody, we studied specific and nonspecific uptake in three breast cancer cell lines: BT-474, SK-BR-3, and MDA-MB-231. Rods exhibited higher specific uptake and lower nonspecific uptake in all cells compared with spheres. This surprising interplay between particle shape and antibodies originates from the unique role of shape in determining binding and unbinding of particles to cell surface. In addition to exhibiting higher binding and internalization, trastuzumab-coated rods also exhibited greater inhibition of BT-474 breast cancer cell growth in vitro to a level that could not be attained by soluble forms of the antibody. The effect of trastuzumab-coated rods on cells was enhanced further by replacing polystyrene particles with pure chemotherapeutic drug nanoparticles of comparable dimensions made from camptothecin. Trastuzumab-coated camptothecin nanoparticles inhibited cell growth at a dose 1,000-fold lower than that required for comparable inhibition of growth using soluble trastuzumab and 10-fold lower than that using BSA-coated camptothecin. These results open unique opportunities for particulate forms of antibodies in therapeutics and diagnostics.

Factors that Control the Circulation Time of Nanoparticles in Blood: Challenges, Solutions and Future Prospects

Numerous types of nanoparticles are being designed for systemic and targeted drug delivery. However, keeping nanoparticles in blood for sufficiently long times so as to allow them to reach their therapeutic target is a major challenge. Upon administration into blood, nanoparticles are quickly opsonized and cleared by the macrophages, thereby limiting their circulation times. Surface-modification of nanoparticles by PEG was developed as the first strategy to prolong nanoparticles circulation. While PEGylation has helped prolong particle circulation, it has several limitations including transient nature of the effect and compromised particle-target interactions. Accordingly, several other approaches have been developed to prolong nanoparticle circulation in blood. These include modification with CD47, modulation of mechanical properties, engineering particle morphology and hitchhiking on red blood cells. In this review, we discuss the factors that affect nanoparticles circulation time and discuss recent progress in development of strategies to prolong circulation time.

Polymer particles that switch shape in response to a stimulus

Particle engineering for biomedical applications has unfolded the roles of attributes such as size, surface chemistry, and shape for modulating particle interactions with cells. Recently, dynamic manipulation of such key properties has gained attention in view of the need to precisely control particle interaction with cells. With increasing recognition of the pivotal role of particle shape in determining their biomedical applications, we report on polymeric particles that are able to switch their shape in real time in a stimulus-responsive manner. The shape-switching behavior was driven by a subtle balance between polymer viscosity and interfacial tension. The balance between the two forces was modulated by application of an external stimulus chosen from temperature, pH, or chemical additives. The dynamics of shape switch was precisely controlled over minutes to days under physiological conditions. Shape-switching particles exhibited unique interactions with cells. Elliptical disk-shaped particles that are not phagocytosed by macrophages were made to internalize through shape switch, demonstrating the ability of shape-switchable particles in modulating interaction with cells.

Endocytosis and Intracellular Distribution of PLGA Particles in Endothelial Cells: Effect of Particle Geometry.

Targeting, internalization, and intracellular trafficking of carriers are key processes in drug delivery to endothelial cells. We synthesized PLGA particles with spherical and elliptical disk geometries and investigated the effect of particle shape on rate of particle endocytosis and their intracellular distribution in endothelial cells. Elliptical disks (aspect ratio of 5) were endocytosed at a slower rate compared to spheres (1.8 µm diameter) of the same volume. However, both particles were eventually internalized and accumulated around the nucleus. We quantified the orientation of elliptical disks and found that disks, on average, oriented tangentially with the nuclear membrane. The non-spherical geometry of elliptical disks brings unique aspects to the kinetics and equilibrium distribution of these particles in cells.

Air-Liquid Interface Culture of Serially Passaged Human Nasal Epithelial Cell Monolayer for In Vitro Drug Transport Studies

The objective of this study was to establish a drug transport study using human nasal epithelial (HNE) cell monolayers cultured by the air-liquid interface (ALI) method using serum-free medium (BEGM:DME/F12, 50:50). The cells were developed and characterized in comparison to those that have been previously cultured by the liquid-covered culture (LCC) method. The epithelial cell monolayer cultured by the ALI method resulted in a significantly higher transepithelial electrical resistance value (3,453 ± 302 ohm × cm2) that was maintained (>1,000 ohm × cm2) for up to 20 days compared with that cultured by the LCC method. Observation by scanning electron microscopy revealed mature cilia after 2 weeks in the ALI culture, while flatten unhealthy ciliated cells were observed in the LCC method. After 21 days, higher level of MUC5AC and 8 mRNA were expressed in ALI culture which confirmed the secretory differentiation of HNE monolayers in vitro. No significant difference in the permeability coefficients of a model hydrophilic marker (14C-mannitol) and a lipophilic drug (budesonide) was observed between the two conditions on day 7. The passage 2–3 of the HNE monolayer using ALI condition retained the morphology and differentiated features of normal epithelium. Thus it would be a suitable model for in vitro nasal drug delivery studies.

Serially Passaged Human Nasal Epithelial Cell Monolayer for in Vitro Drug Transport Studies

AbstractPurpose. To evaluate the feasibility of using a serially passaged culture of human nasal epithelial cell monolayers on a permeable support for in vitro drug transport studies. The optimum conditions for passaged culture as well as the correlation between the transepithelial electrical resistance (TEER) value and drug permeability (Papp) were evaluated. Methods. Fresh human nasal epithelial cells were collected from normal inferior turbinates and were subcultured repeatedly in serum-free bronchial epithelial cell growth media (BEGM) in petri dishes. The subcultured cells of each passage were seeded onto permeable supports at 5 × 105 cells/cm2 and grown in Dulbeccos modified Eagle medium (DMEM). Morphologic characteristics were observed by scanning electron microscopy (SEM). To verify the formation of tight junctions, actin staining and transmission electron microscopy (TEM) studies were conducted. In the drug transport study, [14C]mannitol and budesonide were selected as the paracellular and the transcellular route markers, respectively. Results. Serially passaged cells were successfully cultured on a permeable support and showed significantly high TEER values up to passage 4. After 14 days of seeding, SEM showed microvilli, and protrusions of cilia and mucin granules were detected by TEM. The paracellular marker [14C]mannitol showed a nearly constant permeability coefficient (Papp) when the TEER value exceeded 500 Ω·cm2 regardless of the passage number. However, as expected, budesonide showed a higher permeability coefficient compared to [14C]mannitol and was less affected by the TEER value. Conclusions. Human nasal epithelial cell monolayers were successfully subcultured on a permeable support up to passage 4. These cell culture methods may be useful in high-throughput screening of in vitro nasal transport studies of various drugs.

Size-controlled biodegradable nanoparticles: Preparation and size-dependent cellular uptake and tumor cell growth inhibition

Biodegradable nanoparticles with diameters below 1000nm are of great interest in the contexts of targeted delivery and imaging. In this study, we prepared PLGA nanoparticles with well-defined sizes of ∼70nm (NP70), ∼100nm (NP100), ∼200nm (NP200), ∼400nm (NP400), ∼600nm (NP600) and ∼1000nm (NP1000) using facile fabrication methods based on a nanoprecipitation and solvent evaporation techniques. The nanoparticles showed a narrow size distribution with high yield. Then the size-controlled biodegradable nanoparticles were used to investigate how particle size at nanoscale affects interactions with tumor cells and macrophages. Interestingly, an opposite size-dependent interaction was observed in the two cells. As particle size gets smaller, cellular uptake increased in tumor cells and decreased in macrophages. We also found that paclitaxel (PTX)-loaded nanoparticles showed a size-dependent inhibition of tumor cell growth and the size-dependency was influenced by cellular uptake and PTX release. The size-controlled biodegradable nanoparticles described in this study would provide a useful means to further elucidate roles of particle size on various biomedical applications.

Nitric oxide-releasing poly(lactic-co-glycolic acid)-polyethylenimine nanoparticles for prolonged nitric oxide release, antibacterial efficacy, and in vivo wound healing activity

Nitric oxide (NO)-releasing nanoparticles (NPs) have emerged as a wound healing enhancer and a novel antibacterial agent that can circumvent antibiotic resistance. However, the NO release from NPs over extended periods of time is still inadequate for clinical application. In this study, we developed NO-releasing poly(lactic-co-glycolic acid)-polyethylenimine (PEI) NPs (NO/PPNPs) composed of poly(lactic-co-glycolic acid) and PEI/diazeniumdiolate (PEI/NONOate) for prolonged NO release, antibacterial efficacy, and wound healing activity. Successful preparation of PEI/NONOate was confirmed by proton nuclear magnetic resonance, Fourier transform infrared spectroscopy, and ultraviolet/visible spectrophotometry. NO/PPNPs were characterized by particle size, surface charge, and NO loading. The NO/PPNPs showed a prolonged NO release profile over 6 days without any burst release. The NO/PPNPs exhibited potent bactericidal efficacy against methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa concentration-dependently and showed the ability to bind on the surface of the bacteria. We also found that the NO released from the NO/PPNPs mediates bactericidal efficacy and is not toxic to healthy fibroblast cells. Furthermore, NO/PPNPs accelerated wound healing and epithelialization in a mouse model of a MRSA-infected wound. Therefore, our results suggest that the NO/PPNPs presented in this study could be a suitable approach for treating wounds and various skin infections.

Enzyme/pH dual sensitive polymeric nanoparticles for targeted drug delivery to the inflamed colon.

Novel nanoparticles whose drug release profiles are controlled by both enzyme and pH were prepared for the colon-specific drug delivery using a polymeric mixture of enzyme-sensitive azo-polyurethane and pH-sensitive Eudragit S100 (ES-Azo.pu). The enzyme/pH dual sensitive nanoparticles were designed to release a drug based on a two-fold approach which specifically aimed to target drug delivery to the inflamed colon while preventing the burst release of drugs in the stomach and small intestine. Single pH-sensitive (ES) and dual sensitive (ES-Azo.pu) nanoparticles were prepared using an oil-in-water emulsion solvent evaporation method and coumarin-6 (C-6) was used as a model drug. The successful formation of ES and ES-azo.pu nanoparticles that have 214 nm and 244 nm in mean particle size, respectively, was confirmed by scanning electron microscopy and qNano. ES nanoparticles showed almost 100% of burst drug release at pH 7.4, whereas ES-Azo.pu nanoparticles prevented the burst drug release at pH 7.4, followed by a sustained release phase thereafter. Furthermore, ES-Azo.pu nanoparticles exhibited enzyme-triggered drug release in the presence of rat cecal contents obtained from a rat model of colitis. An in vivo localization study in rat gastrointestinal tract demonstrated that ES-Azo.pu nanoparticles were selectively distributed in the inflamed colon, showing 5.5-fold higher C-6 than ES nanoparticles. In conclusion, the enzyme/pH dual sensitive nanoparticles presented in this study can serve as a promising strategy for colon-specific drug delivery against inflammatory bowel disease and other colon disorders.