Encapsulation of drugs within nanocarriers that selectively target malignant cells promises to mitigate side effects of conventional chemotherapy and to enable delivery of the unique drug combinations needed for personalized medicine. To realize this potential, however, targeted nanocarriers must simultaneously overcome multiple challenges, including specificity, stability, and a high capacity for disparate cargos. Here we report porous nanoparticle-supported lipid bilayers (protocells) that synergistically combine properties of liposomes and nanoporous particles. Protocells modified with a targeting peptide that binds to human hepatocellular carcinoma (HCC) exhibit a 10,000-fold greater affinity for HCC than for hepatocytes, endothelial cells, and immune cells. Furthermore, protocells can be loaded with combinations of therapeutic (drugs, siRNA, and toxins) and diagnostic (quantum dots) agents and modified to promote endosomal escape and nuclear accumulation of selected cargos. The enormous capacity of the high-surface-area nanoporous core combined with the enhanced targeting efficacy enabled by the fluid supported lipid bilayer allow a single protocell loaded with a drug cocktail to kill a drug-resistant HCC cell, representing a 106-fold improvement over comparable liposomes.

We have developed structure/toxicity relationships for amorphous silica nanoparticles (NPs) synthesized through low temperature, colloidal (e.g. Stöber silica) or high temperature pyrolysis (e.g. fumed silica) routes. Through combined spectroscopic and physical analyses, we have determined the state of aggregation, hydroxyl concentration, relative proportion of strained and unstrained siloxane rings, and potential to generate hydroxyl radicals for Stöber and fumed silica NPs with comparable primary particle sizes (16-nm in diameter). Based on erythrocyte hemolytic assays and assessment of the viability and ATP levels in epithelial and macrophage cells, we discovered for fumed silica an important toxicity relationship to post-synthesis thermal annealing or environmental exposure, whereas colloidal silicas were essentially non-toxic under identical treatment conditions. Specifically, we find for fumed silica a positive correlation of toxicity with hydroxyl concentration and its potential to generate reactive oxygen species (ROS) and cause red blood cell hemolysis. We propose fumed silica toxicity stems from its intrinsic population of strained three-membered rings (3MRs) along with its chain-like aggregation and hydroxyl content. Hydrogen-bonding and electrostatic interactions of the silanol surfaces of fumed silica aggregates with the extracellular plasma membrane cause membrane perturbations sensed by the Nalp3 inflammasome, whose subsequent activation leads to secretion of the cytokine IL-1β. Hydroxyl radicals generated by the strained 3MRs in fumed silica but largely absent in colloidal silicas may contribute to the inflammasome activation. Formation of colloidal silica into aggregates mimicking those of fumed silica had no effect on cell viability or hemolysis. This study emphasizes that not all amorphous silica is created equal and that the unusual toxicity of fumed silica compared to colloidal silica derives from its framework and surface chemistry along with its fused chain-like morphology established by high temperature synthesis (>1300°C) and rapid thermal quenching.

Mixing liposomes with hydrophilic particles will induce fusion of the liposome onto the particle surface. Such supported bilayers have been extensively studied as a model for the cell membrane, while its application in drug delivery has not been pursued. In this communication, we report the use of phospholipids to achieve synergistic loading and encapsulating of a fluorescent dye (calcein) in mesoporous silica nanoparticles, and its delivery into mammalian cells. We found that cationic lipid DOTAP provides the highest calcein loading with the concentration inside silica ∼110× higher than that in the solution under experimental conditions. Compared to some other nanoparticle systems, protocells provide a simple construct for loading, sealing, delivering and releasing, and should serve as a useful system in nanomedicine.One of the major challenges in nanomedicine is to engineer nanostructures and materials that can efficiently encapsulate drugs at high concentration, cross the cell membrane, and controllably release the cargo at the target site over a prescribed period of time. 1 Recently, inorganic nanoparticles, including gold, silica, and carbon nanotubes have emerged as a new generation of drug/therapy delivery vehicles in nanomedicine. 2,3 Mesoporous silica nanoparticles are particularly attractive in this regard, because of their biocompatibility and their precisely defined nanoporosity. 4-7 With uniform, tunable pore diameters, ranging from ∼2-5-nm and surfaces areas of 700-1500 m 2 /g, drugs and other components can be loaded by adsorption or capillary filling, and the release profiles adjusted by the combination of pore size and pore surface chemistry. 8 Very recently, elegant gating methods, employing coumarin, 9 azobenzene, 10,11 rotaxane, 12 polymers, 13,14 or small nanoparticles, 15,16 have been established to seal the cargo within the particle and allow its triggered release according to an optical or electrochemical stimulus. Here we describe a synergistic system where liposome fusion on a mesoporous silica particle core simultaneously loads and seals the cargo, creating a 'protocell' construct useful for delivery across the cell membrane (Fig. 1B). We observe that fusion of a positively charged liposome on a negatively charged mesoporous silica core serves to load the core with a negatively charged dye (excluded from the mesopores without lipid) to concentrations that can exceed 100x those in solution. Sealed within the protocell, this membrane impermeable dye can be transported across the cell membrane and slowly released within the cell. Compared to other nanoparticle delivery systems, the protocell is simple and takes advantage of the low toxicity and immunogenicity of liposomes along with their ability to be PEGylated or conjugated to extend circulation time and effect targeting. Compared to liposomes, however, the protocell is more stable and takes advantage of the mesoporous core to control both loading and release. As noted in many other relevant papers the mesoporous E-mail: cjbrink...

The loading and containment of cargo within nanoparticles and their efficient delivery to cells represent a primary challenge in nanomedicine. We report lipid exchange between free and mesoporous silica nanoparticle-supported lipid bilayers as an effective means of containing cargo, controlling charge, and directing delivery to mammalian cells. The delivery of a membrane-impermeable dye (calcein) and a chemotherapeutic drug (doxorubicin) are demonstrated. Exchanged lipid bilayers minimized premature drug release, and an overall positive charge on the supported lipid bilayer effected enhanced delivery.