Targeted drug delivery in cancer cells with red-light photoactivated mesoporous silica nanoparticles.

Mesoporous nanoparticles for drug delivery would benefit significantly from further improvements in targeting efficiency and endosomal release. We present a system based on colloidal mesoporous silica nanoparticles with targeting-ligands and a red-light photosensitizer. This nanoparticle system provides spatial and temporal control of the release of drugs into the cytosol of cancer cells. Furthermore, the system presents a general platform since it can be loaded with different cargos and adapted for targeting of multiple cell types.

"Liquid-phase calcination" of colloidal mesoporous silica nanoparticles in high-boiling solvents.

We report on a novel high temperature liquid phase "calcination" method with trioctylphosphine oxide (TOPO), tri-n-octylamine (TOA), and squalene for removing the template and strengthening the silica network in colloidal mesoporous silica (CMS) nanoparticles. For such materials, the common calcination procedure in air would result in strong agglomeration, thus preventing their use in colloidal suspensions. The highest efficiency of the new approach is obtained by thermal calcination in TOPO at only 275 °C, as shown by an increasing degree of silica condensation, and the retention of the high colloidal stability of the CMS nanoparticles. Moreover, we also show the ability of the TOPO treatment to remove the template, thus saving a preparation step. The resulting CMS nanoparticles retain the ordered mesostructure, high porosity, and large surface area of the original mesoporous nanoparticles, while showing a much greater degree of silica condensation and high stability. The concept of "liquid calcination" represents a powerful general approach for the preparation of stable colloidal porous nanoparticles.

Organ-restricted vascular delivery of nanoparticles for lung cancer therapy.

Nanoparticle-based targeted drug delivery holds promise for treatment of cancers. However, most approaches fail to be translated into clinical success due to ineffective tumor targeting in vivo. Here, the delivery potential of mesoporous silica nanoparticles (MSN) functionalized with targeting ligands for EGFR and CCR2 is explored in lung tumors. The addition of active targeting ligands on MSNs enhances their uptake in vitro but fails to promote specific delivery to tumors in vivo, when administered systemically via the blood or locally to the lung into immunocompetent murine lung cancer models. Ineffective tumor targeting is due to efficient clearance of the MSNs by the phagocytic cells of the liver, spleen, and lung. These limitations, however, are successfully overcome using a novel organ-restricted vascular delivery (ORVD) approach. ORVD in isolated and perfused mouse lungs of Kras-mutant mice enables effective nanoparticle extravasation from the tumor vasculature into the core of solid lung tumors. In this study, ORVD promotes tumor cell-specific uptake of nanoparticles at cellular resolution independent of their functionalization with targeting ligands. Organ-restricted vascular delivery thus opens new avenues for optimized nanoparticles for lung cancer therapy and may have broad applications for other vascularized tumor types.

Immune response to functionalized mesoporous silica nanoparticles for targeted drug delivery.

Multifunctional mesoporous silica nanoparticles (MSN) have attracted substantial attention with regard to their high potential for targeted drug delivery. For future clinical applications it is crucial to address safety concerns and understand the potential immunotoxicity of these nanoparticles. In this study, we assess the biocompatibility and functionality of multifunctional MSN in freshly isolated, primary murine immune cells. We show that the functionalized silica nanoparticles are rapidly and efficiently taken up into the endosomal compartment by specialized antigen-presenting cells such as dendritic cells. The silica nanoparticles showed a favorable toxicity profile and did not affect the viability of primary immune cells from the spleen in relevant concentrations. Cargo-free MSN induced only very low immune responses in primary cells as determined by surface expression of activation markers and release of pro-inflammatory cytokines such as Interleukin-6, -12 and -1ß. In contrast, when surface-functionalized MSN with a pH-responsive polymer capping were loaded with an immune-activating drug, the synthetic Toll-like receptor 7 agonist R848, a strong immune response was provoked. We thus demonstrate that MSN represent an efficient drug delivery vehicle to primary immune cells that is both non-toxic and non-inflammagenic, which is a prerequisite for the use of these particles in biomedical applications.

Genetically designed biomolecular capping system for mesoporous silica nanoparticles enables receptor-mediated cell uptake and controlled drug release.

Effective and controlled drug delivery systems with on-demand release and targeting abilities have received enormous attention for biomedical applications. Here, we describe a novel enzyme-based cap system for mesoporous silica nanoparticles (MSNs) that is directly combined with a targeting ligand via bio-orthogonal click chemistry. The capping system is based on the pH-responsive binding of an aryl-sulfonamide-functionalized MSN and the enzyme carbonic anhydrase (CA). An unnatural amino acid (UAA) containing a norbornene moiety was genetically incorporated into CA. This UAA allowed for the site-specific bio-orthogonal attachment of even very sensitive targeting ligands such as folic acid and anandamide. This leads to specific receptor-mediated cell and stem cell uptake. We demonstrate the successful delivery and release of the chemotherapeutic agent Actinomycin D to KB cells. This novel nanocarrier concept provides a promising platform for the development of precisely controllable and highly modular theranostic systems.

Protease-mediated release of chemotherapeutics from mesoporous silica nanoparticles to ex vivo human and mouse lung tumors.

Nanoparticles allow for controlled and targeted drug delivery to diseased tissues and therefore bypass systemic side effects. Spatiotemporal control of drug release can be achieved by nanocarriers that respond to elevated levels of disease-specific enzymes. For example, matrix metalloproteinase 9 (MMP9) is overexpressed in tumors, is known to enhance the metastatic potency of malignant cells, and has been associated with poor prognosis of lung cancer. Here, we report the synthesis of mesoporous silica nanoparticles (MSNs) tightly capped by avidin molecules via MMP9 sequence-specific linkers to allow for site-selective drug delivery in high-expressing MMP9 tumor areas. We provide proof-of-concept evidence for successful MMP9-triggered drug release from MSNs in human tumor cells and in mouse and human lung tumors using the novel technology of ex vivo 3D lung tissue cultures. This technique allows for translational testing of drug delivery strategies in diseased mouse and human tissue. Using this method we show MMP9-mediated release of cisplatin, which induced apoptotic cell death only in lung tumor regions of Kras mutant mice, without causing toxicity in tumor-free areas or in healthy mice. The MMP9-responsive nanoparticles also allowed for effective combinatorial drug delivery of cisplatin and proteasome inhibitor bortezomib, which had a synergistic effect on the (therapeutic) efficiency. Importantly, we demonstrate the feasibility of MMP9-controlled drug release in human lung tumors.

A fast analysis method to quantify nanoparticle uptake on a single cell level.

AIM: This study examines the absolute quantification of particle uptake into cells. METHODS: We developed a novel method to analyze stacks of confocal fluorescence images of single cells interacting with nano-and micro-particles. Particle_in_Cell-3D is a freely available ImageJ macro. During the image analysis routine, single cells are reconstructed in 3D and split into two volumes - intracellular and the membrane region. Next, particles are localized and color-coded accordingly. The mean intensity of single particles, measured in calibration experiments, is used to determine the absolute number of particles. RESULTS: Particle_in_Cell-3D was successfully applied to measure the uptake of 80-nm mesoporous silica nanoparticles into HeLa cells. Furthermore, it was used to quantify the absolute number of 100-nm polystyrene nanoparticles forming agglomerates of up to five particles; the accuracy of these results was confirmed by super-resolution, stimulated emission depletion microscopy. CONCLUSION: Particle_in_Cell-3D is a fast and accurate method that allows the quantification of particle uptake into cells.