The proton sponge hypothesis: Fable or fact? Vermeulen, Lotte M.P.; De Smedt, Stefaan C.; Remaut, Katrien ...
European journal of pharmaceutics and biopharmaceutics,
August 2018, 2018-Aug, 2018-08-00, Letnik:
129
Journal Article
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In non-viral gene therapy, cationic polymers and lipids are frequently used to encapsulate macromolecular therapeutics into nanoparticles. During their journey to deliver the cargo to the intended ...intracellular target, many biological barriers need to be overcome. One of the major bottlenecks for efficient transfection is the endosomal barrier since nanoparticles often remain entrapped inside endosomes and are trafficked towards the lysosomes where the cargo is degraded. For cationic polymers, the proton sponge hypothesis was introduced in the late ‘90s as a way to explain their endosomal escape properties. However, to date, no consensus has been reached in the scientific community about the validity of this hypothesis due to many contradictory reports. Here we review the sometimes conflicting reports that have been published on the proton sponge hypothesis. We also discuss membrane destabilization and polymer swelling as additional factors that might influence endosomal escape of polyplexes. Based on the key publications on this subject, we aim to launch a consensus on the role of the proton sponge hypothesis in endosomal escape.
The globalization of drug trade leads to the expansion of pharmaceutical counterfeiting. The immense threat of low quality drugs to millions of patients is considered to be an under‐addressed global ...health challenge. Analytical authentication technologies are the most effective methods to identify active pharmaceutical ingredients and impurities. However, most of these analytical testing techniques are expensive and need skilled personnel. To combat counterfeiting of drugs, the package of an increasing number of drugs is being protected through advanced package labeling technologies. Though, package labeling is only effective if the drugs are not repackaged. Therefore “in‐drug labeling,” instead of “drug package labeling,” may become powerful tools to protect drugs. This review aims to overview how advanced micro‐ and nanomaterials might become interesting markers for the labeling of tablets and capsules. Clearly, how well such identifiers can be integrated into “solid drugs” without compromising drug safety and efficacy remains a challenge. Also, incorporation of tags has so far only been reported for the protection of solid drug dosage forms. No doubts that in‐drug labeling technologies for “liquid drugs,” like injectables which contain expensive peptides, monoclonal antibodies, vaccines, dermal fillers, could help to protect them from counterfeiting as well.
The immense threat of low‐quality drugs to millions of patients is considered to be an under‐addressed global health challenge. In addition to analytical authentication technologies, “in‐drug labeling,” including printing or encrypting, as well as microscopic, nanoscopic, and molecular tags, may become powerful tools to protect drugs from counterfeiting.
In gene therapy, endosomal escape represents a major bottleneck since nanoparticles often remain entrapped inside endosomes and are trafficked toward the lysosomes for degradation. A detailed ...understanding of the endosomal barrier would be beneficial for developing rational strategies to improve transfection and endosomal escape. By visualizing individual endosomal escape events in live cells, we obtain insight into mechanistic factors that influence proton sponge-based endosomal escape. In a comparative study, we found that HeLa cells treated with JetPEI/pDNA polyplexes have a 3.5-fold increased endosomal escape frequency compared to ARPE-19 cells. We found that endosomal size has a major impact on the escape capacity. The smaller HeLa endosomes are more easily ruptured by the proton sponge effect than the larger ARPE-19 endosomes, a finding supported by a mathematical model based on the underlying physical principles. Still, it remains intriguing that even in the small HeLa endosomes, <10% of the polyplex-containing endosomes show endosomal escape. Further experiments revealed that the membrane of polyplex-containing endosomes becomes leaky to small compounds, preventing effective buildup of osmotic pressure, which in turn prevents endosomal rupture. Analysis of H1299 and A549 cells revealed that endosomal size determines endosomal escape efficiency when cells have comparable membrane leakiness. However, at high levels of membrane leakiness, buildup of osmotic pressure is no longer possible, regardless of endosomal size. Based on our findings that both endosomal size and membrane leakiness have a high impact on proton sponge-based endosomal rupture, we provide important clues toward further improvement of this escape strategy.
•Endosomal escape is a major bottleneck in cytosolic delivery of nanomaterials.•Endosomal escape mechanisms are divided in pore formation, rupture and membrane fusion.•Several assays are used to ...study endosomal escape mechanisms and efficiency.•The applications and limitations of each assay should be taken into account.•Reliable quantification is necessary to enhance overcoming the endosomal escape barrier.
Successful cytosolic delivery of nanomaterials is becoming more and more important, given the increase in intracellular applications of quantum dots, gold nanoparticles, liposomal drug formulations and polymeric gene delivery vectors. Most nanomaterials are taken up by the cell via endocytosis, yet endosomal escape has long been recognized as a major bottleneck in cytosolic delivery. Although it is essential to detect and reliably quantify endosomal escape, no consensus has been reached so far on the methods to do so. This review will summarize and discuss for the first time the different assays used to investigate this elusive step to date.
Intraperitoneal (IP) drug delivery represents an attractive strategy for the local treatment of peritoneal carcinomatosis (PC). Over the past decade, a lot of effort has been put both in the academia ...and clinic in developing IP therapeutic approaches that maximize local efficacy while limiting systemic side effects. Also nanomedicines are under investigation for the treatment of tumors confined to the peritoneal cavity, due to their potential to increase the peritoneal retention and to target drugs to the tumor sites as compared to free drugs. Despite the progress reported by multiple clinical studies, there are no FDA approved drugs or formulations for specific use in the IP cavity yet. This review discusses the current clinical management of PC, as well as recent advances in nanomedicine-based IP delivery. We address important challenges to be overcome towards designing optimal nanocarriers for IP therapy in vivo.
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In myopia, diabetes and ageing, fibrous vitreous liquefaction and degeneration is associated with the formation of opacities inside the vitreous body that cast shadows on the retina, appearing as ...'floaters' to the patient. Vitreous opacities degrade contrast sensitivity function and can cause notable impairment in vision-related quality of life. Here we introduce 'nanobubble ablation' for safe destruction of vitreous opacities. Following intravitreal injection, hyaluronic acid-coated gold nanoparticles and indocyanine green, which is widely used as a dye in vitreoretinal surgery, spontaneously accumulate on collagenous vitreous opacities in the eyes of rabbits. Applying nanosecond laser pulses generates vapour nanobubbles that mechanically destroy the opacities in rabbit eyes and in patient specimens. Nanobubble ablation might offer a safe and efficient treatment to millions of patients suffering from debilitating vitreous opacities and paves the way for a highly safe use of pulsed lasers in the posterior segment of the eye.
Topical vaginal sustained delivery of siRNA presents a significant challenge due to the short residence time of formulations. Therefore, a drug delivery system capable to adhere to the vaginal mucosa ...is desirable, as it could allow a prolonged delivery and increase the effectiveness of the therapy. The aim of this project is to develop a polymeric solid mucoadhesive system, loaded with lipoplexes, able to be progressively rehydrated by the vaginal fluids to form a hydrogel and to deliver siRNA to vaginal tissues.
To minimize adhesive interactions with vaginal mucus components, lipoplexes were coated with different derivatives of polyethylene glycol: DPSE-PEG2000, DPSE-PEG750 and ceramide-PEG2000. Based on stability and diffusion properties in simulated vaginal fluids, lipoplexes containing DSPE-PEG2000 were selected and incorporated in hydroxyethyl cellulose (HEC) hydrogels. Solid systems, called sponges, were then obtained by freeze-drying. Sponges meet acceptable mechanical characteristics and their hardness, deformability and mucoadhesive properties are not influenced by the presence of lipoplexes. Finally, mobility and stability of lipoplexes inside sponges rehydrated with vaginal mucus, mimicking in situ conditions, were evaluated by advanced fluorescence microscopy. The release rate was found to be influenced by the HEC concentration and consequently by the viscosity after rehydration.
This study demonstrates the feasibility of entrapping pegylated lipoplexes into a solid matrix system for a prolonged delivery of siRNA into the vagina.
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Autophagy or ‘self-eating’ is a process by which defective organelles and foreign material can be cleared from the cell's cytoplasm and delivered to the lysosomes in which degradation occurs. It ...remains an open question, however, whether nanoparticles that did not enter the cell through endocytosis can also be captured from the cytoplasm by autophagy. We demonstrate that nanoparticles that are introduced directly in the cytoplasm of the cells by microinjection, can trigger an autophagy response. Moreover, both polystyrene beads and plasmid DNA containing poly-ethylene-imine complexes colocalize with autophagosomes and lysosomes, as was confirmed by electron microscopy. This indicates that cytoplasmic capturing of nanoparticles can occur by an autophagy response. The capturing of nanoparticles from the cytoplasm most likely limits the time frame in which efficient nucleic acid delivery can be obtained. Hence, autophagy forms an additional barrier to non-viral gene delivery, a notion that was not often taken into account before. Furthermore, these findings urge us to reconsider the idea that a single endosomal escape event is sufficient to have the long-lasting presence of nanoparticles in the cytoplasm of the cells.
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Retinal gene delivery via intravitreal injection is hampered by various physiological barriers present in the eye of which the vitreoretinal (VR) interface represents the most serious hurdle. In this ...study, we present a retinal explant model especially designed to study the role of this interface as a barrier for the penetration of vectors into the retina. In contrast to all existing explant models, the developed model is bovine-derived and more importantly, keeps the vitreous attached to the retina at all times to guarantee an intact VR interface. After ex vivo intravitreal injection into the living retinal explant, the route of fluorescent carriers across the VR interface can be tracked. By applying two different imaging methods on this model, we discovered that the transfer through the VR barrier is size-dependent since 40 nm polystyrene particles are more easily taken up in the retina than 100 and 200 nm sized particles. In addition, we found that removing the vitreous, as commonly done for culture of conventional explants, leads to an overestimation of particle uptake, and conclude that the ultimate barrier to overcome for retinal uptake is undoubtedly the inner limiting membrane. Damaging this matrix resulted in a massive increase in particle transfer into the retina. In conclusion, we have developed a highly relevant ex vivo model that maximally mimics the human in vivo physiology which can be applied as a representative test set-up to assess the potential of promising drug delivery carriers to cross the VR interface.
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