Our project is based on vesicular nanocontainers (NC) consisting of PMOXA-PDMS-PMOXA triblock copolymer building blocks designed for secure content encapsulation. The chosen polymer is biocompatible, impermeable, inert, functionizable, and allows the integration of functional membrane proteins. The hollow NCs have a mean diameter of 200 nm, a wall thickness of 10 nm and a mean volume of 50×10-19 l. By coupling the macrophage scavenger receptor A1 (SRA1) ligand polyguanylic acid to the nanocontainers, we created a receptor-specific drug delivery platform.

Preparation of nanocontainers for receptor targeting: NC’s containing 10% biotinylated copolymer were incubated with streptavidin solution, were separated chromatographically, and were incubated with the biotinylated ligand.

Scanning electron microscopy (SEM) micrograph of nanocontainers shows roughly spherical objects with sizes ranging from 100-250nm diameter. Note that the drying step necessary for SEM might somewhat alter the spherical appearance of nanocontainers as seen in reality.

Targeting specific cells or specific membrane proteins in the body is desirable for improved diagnosis and treatment. Current targeting strategies suffer from significant unspecific uptake in multiple organs or from rapid clearing of the marker from the circulation by renal filtering. Another problem is rapid removal from the circulation of unstable vehicles like liposomes, or the uptake of the substances by the reticuloendothelial system facilitated by unspecific protein binding.

Specific targeting of cells or receptors is important because of undesirable site effects of many drugs. So, cytostatics affect other cell lines (e.g., the fast-growing cells of the bone marrow, immune system, intestinal endothelium, and hair follicles), which limits their therapeutic efficacy. Even more important may be the possibility of delivering therapeutic genes to specific cellular targets within the body.

We explore the potential of synthetic, functional nanocontainers as generic, versatile carriers by examining their interaction with a clinically important, receptor-specific cell model. We focus on design requirements for receptor- and cell-specific binding, on specificity of binding, on the process of surface adherence, and the mode and time course of uptake, as well as on assessment of cytotoxicity. We use macrophages and their scavenger receptor A1 (SRA1) as model target because these cells play a major role in a wide range of pathologies including infection, autoimmunity, cancer, and atherosclerosis.

Polymer nanocontainers seen by transmission (left) and scanning electron microscopy (right)

Main cell types we are working with are human THP-1 macrophages (left) and simian COS-7 cells

Transgenic COS-7 cells expressing the target receptor scavenger receptor A1 (SRA-1) were treated with nanocontainers coupled to the tracking ligand PolyG. Analysis was done by confocal microscopy (FITC filter set left, Texas Red filter set right). The left picture shows the localization of the cellular SRA-1 receptor, which were labeled by co-expression of enhanced green fluorescent protein (EGFP). The right picture shows the localization of the nanocontainers, which were filled with a red fluorescent dye. The pictures demonstrate the receptor-ligand co-localization, an important condition for receptor-selective targeting of nanocontainers. Furthermore, it is evident that the target cell stores the endocytosed nanocontainers in intracellular vesicles - most probably lysosomes. (click here for further details)

Scavenger receptor A1-expressing macrophages derived from human THP-1 cells were treated with fluorescent, ligand-labeled nanocontainers. Analysis by fluorscent microscopy (Dapi filter left, Texas Red filter right) shows the localization of the cells by weak autofluorescence (left). The localization of the nanocontainers is shown by the red fluorescent content (right). The cytosol of the cell is filled with vesicular accumulations of fluorescence, while the cell nucleus shows no signal.