Many emerging therapies rely on the delivery of biological cargo into the cytosol. Nanoparticle delivery systems hold great potential to deliver these therapeutics but are hindered by entrapment and subsequent degradation in acidic compartments of the endo/lysosomal pathway. Engineering polymeric delivery systems that are able to escape the endosome has significant potential to address this issue. However, the development of safe and effective delivery systems that can reliably deliver cargo to the cytosol is still a challenge. Greater understanding of the properties that govern endosomal escape and how it can be quantified is important for the development of more efficient nanoparticle delivery systems. This Topical Review highlights the current understanding of the mechanisms by which nanoparticles escape the endosome, and the emerging techniques to improve the quantification of endosomal escape.

Using nanoparticles to deliver drugs to cells has the potential to revolutionize the treatment of many diseases, including HIV, cancer, and diabetes. One of the major challenges facing this field is controlling where the drug is trafficked once the nanoparticle is taken up into the cell. In particular, if drugs remain localized in an endosomal or lysosomal compartment, the therapeutic can be rendered completely ineffective. To ensure the design of more effective delivery systems we must first develop a better understanding of how nanoparticles and their cargo are trafficked inside cells. This needs to be combined with an understanding of what characteristics are required for nanoparticles to achieve endosomal escape, along with methods to detect endosomal escape effectively. This review is focused into three sections: first, an introduction to the mechanisms governing internalization and trafficking in cells, second, a discussion of methods to detect endosomal escape, and finally, recent advances in controlling endosomal escape from polymer- and lipid-based nanoparticles, with a focus on engineering materials to promote endosomal escape. WIREs Nanomed Nanobiotechnol 2017, 9:e1452. doi: 10.1002/wnan.1452 For further resources related to this article, please visit the WIREs website.

The ability to determine the amount of material endocytosed by a cell is important for our understanding of cell biology and in the design of effective carriers for drug delivery. To quantify internalization by fluorescence, the signal from material remaining on the cell surface must be differentiated from endocytosed material. Sensors for internalization offer advantages over traditional methods for achieving this as they exhibit improved sensitivity, allow for multiple fluorescent markers to be used simultaneously, and are amenable to high-throughput analysis. We have developed a small fluorescent internalization sensor, similar in size to a standard fluorescent dye, that can be conjugated to proteins and uses the rapid and highly specific bio-orthogonal reaction between a tetrazine and a trans-cyclooctene group to switch off the surface signal. The sensor can be attached to a variety of materials using simple chemistry and is compatible with flow cytometry and fluorescence microscopy, making it a useful tool to study the uptake of material into cells.

SHIP is able to distinguish between nanoparticles that are associated on the outer cell membrane from nanoparticles that are internalized. This study demonstrates the assay can be used to probe the kinetics of nanoparticle internalization and the mechanisms by which the nanoparticles are taken up by cells. This information is fundamental for engineering more effective nanoparticle delivery systems. The SHIP assay is a simple and a high-throughput technique that could have wide application in therapeutic delivery research.

Targeting nanoparticles to specific cellular receptors has the potential to deliver therapeutic compounds to target sites while minimizing side effects. To this end, we have conjugated a targeting protein, holo-transferrin (holo-Tf), to pH-responsive polymers, poly­(2-(diethylamino)­ethyl methacrylate) (PDEAEMA) and poly­(2-(diethylamino)­ethyl methacrylate)-ran-poly­(2-(diisopropylamino)­ethyl methacrylate (PDEAEMA-r-PDPAEMA). These protein–polymer hybrid materials were observed to self-assemble when the pH is increased above the pK a of the polymer. We demonstrate that their response to pH could be tuned depending on the polymer constituent attached to holo-Tf. Importantly, the targeting behavior of these nanoparticles could be maximized by tuning the density of holo-Tf on the nanoparticle surface by the introduction of a (PDEAEMA-r-PDPAEMA)-b-poly­(ethylene glycol) (PEG) copolymer.