Metal-organic frameworks as intracellular delivery vectors

Metal-organic frameworks (MOFs) have emerged as interesting candidates for intracellular carrier-based delivery. These hybrid materials are constituted of metal clusters linked together by organic ligands. The possibility to tune their physical and chemical properties both in the bulk and at the surface allows for the design of biocompatible delivery systems with high loading capacities and targeting abilities, combining the benefits of both organic and inorganic materials. The following dissertation focuses on developing and evaluating MOFs as intracellular delivery systems. In the first instance, a zirconium-based MOF, UiO-66, was synthesised and utilised as an intracellular delivery vector for trehalose, a disaccharide with cryoprotective properties when present in the cytosol. This MOF demonstrated very high trehalose weight loadings compared to other trehalose delivery systems (up to ca. 50 wt %), release of the sugar from the framework over 5 h, and appropriate biocompatibility. To assess the delivery system’s impact on cryopreservation, the viability of cells cryoprotected with trehalose-loaded UiO-66 was tested at 0 h, 24 h, and 48 h post-thaw, and showed no improvement compared to cells frozen with free trehalose or growth media alone. The absence of cryoprotective effect was hypothesised to be due to endosomal entrapment of the delivery system after cellular uptake through endocytosis. The final fate of particles taken up by cells depends on the endocytosis pathways they go through. In order to confirm the hypothesis of MOF endosomal entrapment, the endocytosis of MOF particles was studied. In particular, the effects of surface chemistry of Zr-based MOFs on their endocytosis mechanisms were investigated. It was found that MOF surface chemistry had an important effect on cellular uptake behaviour, whereas particle size played a less important role. In particular, Zr-based MOFs synthesised using naphthalene-2,6-dicarboxylic acid and 4,4′-biphenyldicarboxylic acid as organic ligands, and UiO 66 particles surface-decorated with folic acid and PEG, promoted entry through the caveolin-pathway. This allowed the particles to potentially avoid endosomal entrapment and reach the cytosol, enhancing their therapeutic activity when loaded with drugs. Equipped with an understanding of the cellular uptake of MOF particles, a range of mitochondrially-targeted UiO-66 particles capable of bypassing endosomal entrapment was prepared and tested. The UiO-66 particles were loaded with dichloracetic acid (DCA), a small chemotherapeutic drug molecule that acts on mitochondria, and surface-functionalised with triphenylphosphonium, a known mitochondrial targeting agent. The system demonstrated a dramatic increase in efficacy, allowing a reduction in DCA effective dose of ca. 100-fold compared to the free drug, and ca. 10-fold compared to non-targeted, DCA-loaded UiO-66. Confocal microscopy revealed a distribution of the targeted nanoparticles around mitochondria. Super-resolution microscopy of cells treated with the system revealed important mitochondrial morphology changes associated with cell death as soon as 30 minutes after incubation. A whole transcriptome analysis of cells treated with the system indicated widespread changes in gene expression compared to both untreated cells and to cells treated with non-targeted, DCA-loaded UiO-66. In summary, these studies demonstrated the advantages of MOFs as targeted intracellular delivery vectors. The ease with which their physicochemical properties can be tuned allows for the design of delivery systems able to bypass the critical drug delivery bottlenecks of endosomal entrapment and non-specific delivery

An Avoidable Cognitive Error in Chest Radiography

Teaching Point: Awareness in radiology reporting of cognitive errors such as the alliterative bias can help minimize the delay to diagnosis and accelerate adequate patient care

Selective Surface PEGylation of UiO-66 Nanoparticles for Enhanced Stability, Cell Uptake, and pH-Responsive Drug Delivery.

The high storage capacities and excellent biocompatibilities of metal-organic frameworks (MOFs) have made them emerging candidates as drug-delivery vectors. Incorporation of surface functionality is a route to enhanced properties, and here we report on a surface-modification procedure-click modulation-that controls their size and surface chemistry. The zirconium terephthalate MOF UiO-66 is (1) synthesized as ∼200 nm nanoparticles coated with functionalized modulators, (2) loaded with cargo, and (3) covalently surface modified with poly(ethylene glycol) (PEG) chains through mild bioconjugate reactions. At pH 7.4, the PEG chains endow the MOF with enhanced stability toward phosphates and overcome the "burst release" phenomenon by blocking interaction with the exterior of the nanoparticles, whereas at pH 5.5, stimuli-responsive drug release is achieved. The mode of cellular internalization is also tuned by nanoparticle surface chemistry, such that PEGylated UiO-66 potentially escapes lysosomal degradation through enhanced caveolae-mediated uptake. This makes it a highly promising vector, as demonstrated for dichloroacetic-acid-loaded materials, which exhibit enhanced cytotoxicity. The versatility of the click modulation protocol will allow a wide range of MOFs to be easily surface functionalized for a number of applications

Tuning the endocytosis mechanism of Zr-based metal−organic frameworks through linker functionalization

A critical bottleneck for the use of metal-organic frameworks (MOFs) as drug delivery systems has been allowing them to reach their intracellular targets without being degraded in the acidic environment of the lysosomes. Cells take up particles by endocytosis through multiple biochemical pathways, and the fate of these particles depends on these routes of entry. Here, we show the effect of functional group incorporation into a series of Zr-based MOFs on their endocytosis mechanisms, allowing us to design an effi-cient drug delivery system. In particular, naphthalene-2,6-dicarboxylic acid and 4,4'-biphenyldicarboxylic acid ligands promote entry through the caveolin-pathway, allowing the particles to avoid lysosomal degradation and be delivered into the cytosol, en-hancing their therapeutic activity when loaded with drugs

Mechanistic investigation into the selective anticancer cytotoxicity and immune system response of surface-functionalised, dichloroacetate-loaded, UiO-66 nanoparticles

The high drug loading and excellent biocompatibilities of metal-organic frameworks (MOFs) have led to their application as drug delivery systems (DDSs). Nanoparticle surface chemistry dominates both biostability and dispersion of DDSs while governing their interactions with biological systems, cellular and/or tissue targeting, and cellular internalisation, leading to a requirement for versatile and reproducible surface functionalisation protocols. Herein, we explore not only the effect of introducing different surface functionality to the biocompatible Zr-MOF UiO-66, but also the efficacy of three surface modification protocols: (i) direct attachment of biomolecules (folic acid, biotin) introduced as modulators of UiO-66 synthetic, (ii) our previously reported ‘’click-modulation” approach to covalently attach polymers (poly(ethylene glycol), poly-L-lactide, poly-N-isopropylacrylamide) to the surface of UiO-66 through click chemistry, and (iii) surface ligand exchange, to postsynthetically coordinate folic acid, biotin and heparin to UiO-66. The innovative use of a small molecule with metabolic anticancer activity, dichloroacetic acid (DCA), as a modulator during synthesis is described, and found to be compatible with all three protocols, yielding surface-coated, DCA-loaded (10-20% w/w) nanoMOFs (70-170 nm). External surface modification generally enhances stability and colloidal dispersion of UiO-66. Cellular internalisation routes and efficiencies of UiO-66 by HeLa cervical cancer cells can be tuned by surface chemistry, and anticancer cytotoxicity of DCA-loaded MOFs correlates with endocytosis efficiency and mechanisms. The MOFs with the most promising coatings (folic acid, poly(ethylene glycol), poly-L-lactide, and poly-N-isopropylacrylamide) were extensively tested for selectivity of anti-cancer cytotoxicity against MCF-7 breast cancer cells and HEK293 healthy kidney cells, as well as for cell proliferation and ROS production against J774 macrophages and peripheral blood lymphocytes (PBLs) isolated from the blood of human donors. DCA-loaded, folic acid modified UiO-66 selectively kills cancer cells without harming healthy ones or provoking immune system response in vitro, suggesting a significant targeting effect and great potential in anticancer drug delivery. The results provide mechanistic insight into the design and functionalisation of MOFs for drug delivery, and underline the availability of various in vitro techniques to potentially minimise early-stage in vivo animal studies, following the three Rs: reduction, refinement and replacement

Surface-functionalisation of Zr-Fumarate MOF for selective cytotoxicity and immune system compatibility in nanoscale drug delivery

Metal-organic frameworks (MOFs), network structures wherein metal ions or clusters link organic ligands into porous materials, are being actively researched as nanoscale drug delivery devices (DDSs) as they offer tuneable structures with high cargo loading that can easily be further functionalized for targeting and enhanced physiological stability. The excellent biocompatibility of Zr has meant that its MOFs are amongst the most studied to date, in particular the archetypal Zr terephthalate UiO-66. In contrast, the isoreticular analogue linked by fumarate (Zr-fum) has received little attention, despite the endogenous linker being part of the Krebs cycle. Herein, we report a comprehensive study of Zr-fum in the context of drug delivery. Reducing particle size is shown to increase uptake by cancer cells while reducing internalisation by macrophages, immune system cells that remove foreign objects from the bloodstream. Zr-fum is compatible with defect-loading of the drug dichloroacetate, as well as surface modification during synthesis, through coordination modulation, and postsynthetically. DCA-loaded, PEGylated Zr-fum shows selective in vitro cytotoxicity towards HeLa and MCF-7 cancer cells, likely as a consequence of its enhanced caveolae-mediated endocytosis compared to uncoated precursors, and it is well tolerated by HEK293 kidney cells, J774 macrophages, and human peripheral blood lymphocytes. Compared to UiO-66, Zr-fum is more efficient at transporting the drug mimic calcein into HeLa cells, and DCA-loaded, PEGylated Zr-fum is more effective at reducing HeLa and MCF-7 cell proliferation than the analogous UiO-66 sample. In vitro examination of immune system response shows Zr-fum samples induce less reactive oxygen species than UiO-66 analogues, possibly as a consequence of the linker being endogenous, and do not activate the C3 and C4 complement cascade pathways, suggesting that Zr-fum can avoid phagocytic activation. The results show that Zr-fum is an attractive alternative to UiO-66 for nanoscale drug delivery, and that a wide range of in vitro experiments are available to greatly inform the design of DDSs prior to early stage animal studies

Design of a Functionalized Metal-Organic Framework System for Enhanced Targeted Delivery to Mitochondria.

Mitochondria play a key role in oncogenesis and constitute one of the most important targets for cancer treatments. Although the most effective way to deliver drugs to mitochondria is by covalently linking them to a lipophilic cation, the in vivo delivery of free drugs still constitutes a critical bottleneck. Herein, we report the design of a mitochondria-targeted metal-organic framework (MOF) that greatly increases the efficacy of a model cancer drug, reducing the required dose to less than 1% compared to the free drug and ca. 10% compared to the nontargeted MOF. The performance of the system is evaluated using a holistic approach ranging from microscopy to transcriptomics. Super-resolution microscopy of MCF-7 cells treated with the targeted MOF system reveals important mitochondrial morphology changes that are clearly associated with cell death as soon as 30 min after incubation. Whole transcriptome analysis of cells indicates widespread changes in gene expression when treated with the MOF system, specifically in biological processes that have a profound effect on cell physiology and that are related to cell death. We show how targeting MOFs toward mitochondria represents a valuable strategy for the development of new drug delivery systems

Back to the future : the Arab uprisings and state (re)formation in the Arab world

This article contributes to debates that aim to go beyond the “democratization” and “post-democratization” paradigms to understand change and continuity in Arab politics. In tune with calls to focus on the actualities of political dynamics, the article shows that the literatures on State Formation and Contentious Politics provide useful theoretical tools to understand change/continuity in Arab politics. It does so by examining the impact of the latest Arab uprisings on state formation trajectories in Iraq and Syria. The uprisings have aggravated a process of regime erosion – which originated in post-colonial state-building attempts – by mobilizing sectarian and ethnic identities and exposing the counties to geo-political rivalries and intervention, giving rise to trans-border movements, such as ISIS. The resulting state fragmentation has obstructed democratic transition in Syria and constrained its consolidation in Iraq.PostprintPeer reviewe

Design of a functionalized metal-organic framework system for enhanced targeted delivery to mitochondria

Mitochondria play a key role in oncogenesis and constitute one of the most important targets for cancer treatments. Although the most effective way to deliver drugs to mitochondria is by covalently linking them to a lipophilic cation, the in vivo delivery of free drugs still constitutes a critical bottleneck. Herein, we report the design of a mitochondria-targeted metal-organic framework (MOF) that greatly increases the efficacy of a model cancer drug, reducing the required dose to less than 1% compared to the free drug and ca. 10% compared to the non-targeted MOF. The performance of the system is evaluated using a holistic approach ranging from microscopy to transcriptomics. Super-resolution microscopy of MCF-7 cells treated with the targeted MOF system reveals important mitochondrial morphology changes that are clearly associated with cell death as soon as 30 minutes after incubation. Whole transcriptome analysis of cells indicated widespread changes in gene expression when treated with the MOF system, specifically in biological processes that have a profound effect on cell physiology and that are related to cell death. We show how targeting MOFs towards mitochondria represents a valuable strategy for the development of new drug delivery systems