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Synthesis of cyclodextrin-based polyrotaxanes and polycatenanes for supramolecular pharmaceutical sciences
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Accumulation of lipotoxic lipids, such as free cholesterol, induces hepatocyte death and subsequent inflammation and fibrosis in the pathogenesis of nonalcoholic steatohepatitis (NASH). However, the underlying mechanisms remain unclear. We have previously reported that hepatocyte death locally induces phenotypic changes in the macrophages surrounding the corpse and remnant lipids, thereby promoting liver fibrosis in a murine model of NASH. Here, we demonstrated that lysosomal cholesterol overload triggers lysosomal dysfunction and profibrotic activation of macrophages during the development of NASH. β-cyclodextrin polyrotaxane (βCD-PRX), a unique supramolecule, is designed to elicit free cholesterol from lysosomes. Treatment with βCD-PRX ameliorated cholesterol accumulation and profibrotic activation of macrophages surrounding dead hepatocytes with cholesterol crystals, thereby suppressing liver fibrosis in a NASH model, without affecting the hepatic cholesterol levels. In vitro experiments revealed that cholesterol-induced lysosomal stress triggered profibrotic activation in macrophages predisposed to the steatotic microenvironment. This study provides evidence that dysregulated cholesterol metabolism in macrophages would be a novel mechanism of NASH.
Activation of autophagy represents a potential therapeutic strategy for the treatment of diseases that are caused by the accumulation of defective proteins and the formation of abnormal organelles. Methylated β-cyclodextrins-threaded polyrotaxane (Me-PRX), a supramolecular structured polymer, induces autophagy by interacting with the endoplasmic reticulum. We previously reported on the successful activation of mitochondria-targeted autophagy by delivering Me-RRX to mitochondria using a MITO-Porter, a mitochondria-targeted nanocarrier. The same level of autophagy induction was achieved at one-twentieth the dosage for the MITO-Porter (Me-PRX) compared to the naked Me-PRX. We report herein on the quantitative evaluation of the intracellular organelle localization of both naked Me-PRX and the MITO-Porter (Me-PRX). Mitochondria, endoplasmic reticulum and lysosomes were selected as target organelles because they would be involved in autophagy induction. In addition, organelle injury and cell viability assays were performed. The results showed that the naked Me-PRX and the MITO-Porter (Me-PRX) were localized in different intracellular organelles, and organelle injury was different, depending on the route of administration, indicating that different organelles contribute to autophagy induction. These findings indicate that the organelle to which the autophagy-inducing molecules are delivered plays an important role in the level of induction of autophagy.
Cyclodextrins (CDs) are one of the most extensively studied cyclic-oligosaccharides due to their low toxicity, good biodegradability and biocompatibility, facile chemical modification, and unique inclusion capacity. However, problems such as poor pharmacokinetics, plasma membrane disruption, hemolytic effects and a lack of target specificity still exist for their applications as drug carriers. Recently, polymers have been introduced into CDs to combine the advantages of both biomaterials for the superior delivery of anticancer agents in cancer treatment. In this review, we summarize four types of CD-based polymeric carriers for the delivery of chemotherapeutics or gene agents for cancer therapy. These CD-based polymers were classified based on their structural properties. Most of the CD-based polymers were amphiphilic with the introduction of hydrophobic/hydrophilic segments and were able to form nanoassemblies. Anticancer drugs could be included in the cavity of CDs, encapsulated in the nanoparticles or conjugated on the CD-based polymers. In addition, the unique structures of CDs enable the functionalization of targeting agents and stimuli-responsive materials to realize the targeting and precise release of anticancer agents. In summary, CD-based polymers are attractive carriers for anticancer agents.
Recently, the potential of β-cyclodextrin-thread acid-degradable polyrotaxane (AdPRX) has been emphasized as a therapeutic agent for cholesterol-related metabolic disorders. In this study, we investigated whether carboxymethyl carbamate-modified AdPRX (CMC-AdPRX) can be used for adsorption to calcium phosphate to treat bone diseases. We first synthesized CMC-AdPRX and used it to coat the calcium phosphate plate. RAW264.7 cells were then differentiated into osteoclasts via a receptor activator of nuclear factor-κB ligand, and the number of osteoclasts and the area of absorption lacunae were determined. The number of tartrate-resistant acid phosphatase-positive multinucleated cells was reduced on the CMC-AdPRX-coated plate. The area of the absorption lacunae was smaller with CMC-AdPRX than with AdPRX, which was not carboxy-modified. Our results suggest that CMC-AdPRX can adsorb to calcium phosphate and act on differentiated osteoclasts to suppress their functional expression.
In recent years, a number of interlocked molecules such as rotaxanes, catenanes, polyrotaxanes, and polycatenanes have been actively synthesized. Cyclodextrins (CDs) are representative building blocks of these interlocked molecules; however, very few reports are available on CD-based catenanes and polycatenanes. In this review article, we provide an overview of CD-based interlocked molecules and introduce examples of CD-based catenanes and polycatenanes. Finally, the perspectives of research on CD-based catenanes and polycatenanes are discussed.
In this work, we report a high-yield one-pot synthesis of polyrotaxane (PR), composed of (2-hydroxypropyl)-α-cyclodextrin (hpCD) and polyethylene glycol (PEG), with well-defined hpCD threading ratios controllable across a wide range from 0.64% to 10%. In hpCD/PEG aqueous solutions, hpCDs are well dispersed and threaded spontaneously into hpCDs to form a pseudo-PR (pPR) structure. The homogeneous dispersion of hpCDs results in a well-defined threading ratio of hpCDs on PEG, which is suggested by the fact that the dispersity of the molecular weight distribution of PR is almost the same as that of pure PEG. The well-defined hpCD threading ratio of the PRs can be controlled over a wide range by tuning the hpCD concentration in the pPR solutions.
In recent years, functional materials using cyclodextrin (CD)-based polyrotaxanes have attracted considerable attention, and derivatization reactions are often performed to develop them because of the insolubility of the parent polyrotaxane in water. Recently, we prepared CD-based radial polycatenanes in one-pot through polypseudorotaxane formation between β-CD and thiolated pluronic and subsequent cyclization via disulfide bonds. However, no reports are available on syntheses of CD polycatenane derivatives. The disulfide bonds are not stable enough to synthesize CD polycatenane derivatives; therefore, we herein prepared a chemically stable β-CD polycatenane via amide bond through cyclization of β-CD/aminated pluronic polypseudorotaxane with terephthaloyl chloride. This allowed the derivatization of the polycatenane owing to its stable chemical structure, and resulted in the syntheses of three different water soluble polycatenane derivatives: aminated, succinylated, and hydroxypropylated polycatenanes. These findings may provide useful information for the fabrication of polycatenane-based materials such as slide-ring materials, drug carriers, and biomaterials.
Supermolecules such as rotaxanes and catenanes have recently attracted considerable attention due to their potential widespread applications in areas such as molecular machines and switches. Moreover, the development of polyrotaxanes and polycatenanes, comprising multiple cyclic compounds, has allowed the fabrication of structures with novel properties. Although rotaxanes and polyrotaxanes have been extensively prepared from cyclodextrins as building blocks, very few studies have considered the syntheses of cyclodextrin-based polycatenanes. Here we report the one-pot syntheses and isolation of cyclodextrin-based radial polycatenanes with large numbers of cyclic components (>10) attached to a poly(ethylene glycol)–poly(propylene glycol)–poly(ethylene glycol) copolymer core, with characterization performed using Raman spectroscopy, gel permeation chromatography, ¹H-NMR spectroscopy, and other techniques. Overall, the results presented herein may be used to develop advanced supramolecular structures and materials, such as molecular machines, molecular actuators, molecular switches, biomaterials, and drug carriers.
Gene editing with clustered regularly interspaced short palindromic repeats and CRISPR‐associated protein 9 (CRISPR/Cas9) has shown promise in models of Duchenne muscular dystrophy (DMD); however, nonviral strategies to deliver CRISPR to muscle have not been widely explored or optimized. Most studies have relied on viral vectors, which are likely limited to single dosing due to their immunogenicity, thus reducing their therapeutic potential. Therefore, there is a need to develop nonviral approaches that allow for delivery and repeat dosing of CRISPR/Cas9 therapies to skeletal muscle. Here, biocompatible multi‐arm polyrotaxane (PRX) nanocarriers, are iteratively optimized for packaging large plasmid DNA for delivery to muscle cells. The PRXs are optimized by addition of a disulfide‐responsive linker that enhances plasmid release. Furthermore, conjugation of peptides leads to quicker uptake and improved transfection efficiency in humanized dystrophic muscle cells in vitro. Finally, in vitro delivery of PRXs complexed with a CRISPR/Cas9 platform demonstrates effective deletion of DMD exons 45–55, a therapeutic strategy with potential to restore the reading frame for half of DMD patients. This work represents the first PRX platform that is optimized and designed for delivery of large plasmid DNA, such as CRISPR/Cas9, to dystrophic muscle cells.
Supramolecular chemistry is an extremely useful and important domain for understanding pharmaceutical sciences because various physiological reactions and drug activities are based on supramolecular chemistry. However, it is not a major domain in the pharmaceutical field. In this review, we propose a new concept in pharmaceutical sciences termed “supramolecular pharmaceutical sciences,” which combines pharmaceutical sciences and supramolecular chemistry. This concept could be useful for developing new ideas, methods, hypotheses, strategies, materials, and mechanisms in pharmaceutical sciences. Herein, we focus on cyclodextrin (CyD)-based supermolecules, because CyDs have been used not only as pharmaceutical excipients or active pharmaceutical ingredients but also as components of supermolecules.
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Niemann-Pick Type C disease (NPC) is a rare metabolic disorder characterized by disruption of normal cholesterol trafficking within the cells of the body. There are no FDA approved treatments available for NPC patients. Recently, the cycloheptaglucoside 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) has shown efficacy as a potential NPC therapeutic by extending lifetime in NPC mice, delaying neurodegeneration, and decreasing visceral and neurological cholesterol burden. Although promising, systemic HP-β-CD treatment is limited by a pharmacokinetic profile characterized by rapid loss through renal filtration. To address these shortcomings, we sought to design a family of HP-β-CD pro-drug delivery vehicles, known as polyrotaxanes (PR), capable of increasing the efficacy of a given injected dose by improving both pharmacokinetic profile and bioavailability of the HP-β-CD agent. PR can effectively diminish the cholesterol pool within the liver, spleen, and kidney at molar concentrations 10-to-100-fold lower than monomeric HP-β-CD. In addition to this proof-of-concept, use of PR scaffolds with differing physiochemical properties reveal structure-activity relationships in which PR characteristics, including hydrophobicity, threading efficiency and surface charge, were found to both decisively and subtly effect therapeutic efficacy. PR scaffolds exhibit absorption, pharmacokinetics, and biodistribution patterns that are significantly altered from monomeric HP-β-CD. In all, PR scaffolds hold great promise as potential treatments for visceral disease in NPC patients.
Niemann-Pick type C (NPC) disease is characterized by the accumulation of cholesterol in lysosomes. We have previously reported that biocleavable polyrotaxanes (PRXs) composed of β-cyclodextrins (β-CDs) threaded onto a linear polymer capped with bulky stopper molecules via intracellularly cleavable linkers show remarkable cholesterol reducing effects in NPC disease patient-derived fibroblasts owing to the stimuli-responsive intracellular dissociation of PRXs and subsequent β-CD release from the PRXs. Herein, we describe a series of novel acid-labile 2-(2-hydroxyethoxy)ethyl group-modified PRXs (HEE-PRXs) bearing terminal N-triphenylmethyl (N-Trt) groups as a cleavable component for the treatment of NPC disease. The N-Trt end groups of the HEE-PRXs underwent acidic pH-induced cleavage and led to the dissociation of their supramolecular structure. A kinetic study revealed that the number of HEE groups on the PRX did not affect the cleavage kinetics of the N-Trt end groups of the HEE-PRXs. The effect of the number of HEE groups of the HEE-PRXs, which was modified to impart water solubility to the PRXs, on cellular internalization efficiency, lysosomal localization efficiency, and cholesterol reduction ability in NPC disease-derived fibroblasts (NPC1 fibroblasts) was also investigated. The cellular uptake and lysosomal localization efficiency were almost equivalent for HEE-PRXs with different numbers of HEE groups. However, the cholesterol reducing ability of the HEE-PRXs in NPC1 fibroblasts was affected by the number of HEE groups, and HEE-PRXs with a high number of HEE groups were unable to reduce lysosomal cholesterol accumulation. This deficiency is most likely due to the cholesterol-solubilizing ability of HEE-modified β-CDs released from the HEE-PRXs. We conclude that the N-Trt group acts as a cleavable component to induce the lysosomal dissociation of HEE-PRXs, and acid-labile HEE-PRXs with an optimal number of HEE groups (4.1 to 5.4 HEE groups per single β-CD threaded onto the PRX) have great therapeutic potential for treating NPC disease.
The bottom-up synthesis of highly complex functional materials from simple modular blocks is an intriguing area of research. Driven by the chemistry of supramolecular assembly, modules which self-assemble into intricate structures have been described. These hierarchically assembled systems extend beyond the individual molecule and rely on non-covalent interactions in a directed self-assembly process. The intrinsic properties of the materials can be modified by exploiting the dynamic and specific uni-directional interactions among the building. This also allows the building of novel supramolecular structures such as hydrogels, micelles and vesicles. These aqueous supramolecular networks belong to a novel category of soft biomaterials exhibiting attractive properties such as stimuli-responsiveness and self-healing properties derived from their dynamic behavior. These are important for a wide variety of emerging applications. In this review, the latest literature describing the formation of dynamic polymeric networks through host-guest complex formation will be summarised. These approaches carried out in the aqueous medium have unlocked a versatile toolbox for the design and fine-tuning of supramolecular self-assembled materials.
Niemann-Pick type C (NPC) disease is an autosomal recessive lysosomal trafficking disorder, in which the cholesterols are abnormally accumulated in lysosomes. Recently, the β-cyclodextrin (CD) derivatives are revealed to show therapeutic effect for NPC disease through the removal of accumulated cholesterols in lysosomes. Herein, to enhance the therapeutic effect and reduce the toxicity of β-CD derivatives, biocleavable Pluronic/β-CD-based polyrotaxanes (PRXs) bearing terminal disulfide linkages that can release threaded β-CDs in lysosomes were developed. The biocleavable PRXs show negligible interaction with the plasma membrane, leading to avoiding the toxicity of β-CDs derived from their hydrophobic cavity. Additionally, lysosomal release of threaded β-CDs from biocleavable PRXs by the intracellular cleavage of terminal disulfide linkages is found to achieve approximately 100-fold higher cholesterol removal ability from NPC disease-derived cells than β-CD derivatives. Consequently, the biocleavable PRXs is considered to be a noninvasive and effective therapeutics for NPC disease.
Various biopharmaceuticals, such as nucleic acids, proteins, and genome-editing molecules, have been developed. Generally, carriers are prepared for each biopharmaceutical to deliver it intracellularly; thus, the applications of individual carriers are limited. Moreover, the development of carriers is laborious and expensive. Therefore, in the present study, versatile and universal delivery carriers were developed for various biopharmaceuticals using aminated polyrotaxane libraries. Step-by-step and logical screening revealed that aminated polyrotaxane, including the carbamate bond between the axile molecule and endcap, is suitable as a backbone polymer. Movable and flexible properties of the amino groups modified on polyrotaxane facilitated efficient complexation with various biopharmaceuticals, such as small interfering RNA, antisense oligonucleotides, messenger RNA, β-galactosidase, and genome-editing ribonucleoproteins. Diethylenetriamine and cystamine modifications of polyrotaxane provided endosomal-escape abilities and drug-release properties in the cytosol, allowing higher delivery efficacies than commercially available high-standard carriers without cytotoxicity. Thus, the resulting polyrotaxane might serve as a versatile and universal delivery platform for various biopharmaceuticals.
Acid-degradable polyrotaxanes (PRXs) containing threading β-cyclodextrins (β-CDs) are promising candidates for therapeutic applications of β-CDs in metabolic diseases with cholesterol overload or imbalance. To improve cellular uptake specificity and efficiency of PRXs in hepatocytes, N-acetyl-d-galactosamine (GalNAc)-modified PRXs were developed to facilitate asialoglycoprotein receptor (ASGR)-mediated endocytosis. Binding affinity studies revealed that the dissociation constant (KD) values between recombinant ASGR and GalNAc-PRXs decreased with an increase in the number of modified GalNAc units. Additionally, the KD values for GalNAc-PRXs were smaller than those for GalNAc-modified β-CD and amylose, suggesting that the PRX backbone structure improves the binding affinity with ASGR. However, the intracellular uptake levels of GalNAc-PRXs in HepG2 cells increased with a decrease in the number of modified GalNAc units, which was opposite to the trend observed in the binding affinity study. We found that GalNAc-PRXs had a large number of GalNAc units localized in recycling endosomes, resulting in the low intracellular uptake. The cholesterol-reducing abilities of GalNAc-PRXs were assessed using cholesterol-overloaded HepG2 cells. GalNAc-PRXs with a small number of GalNAc units were demonstrated to show superior cholesterol-reducing effects compared to previously designed acid-degradable PRX and clinically tested β-CD derivatives. Thus, we conclude that GalNAc modification is a promising molecular design for the therapeutic application of β-CD-threaded PRXs in various metabolic diseases with cholesterol overload or imbalance in the liver.
To achieve a systemic targeted delivery of siRNA using polymeric carriers, there is a dilemma between ligand modification and stabilization of the polyplex. Namely, ligand modification often leads to destabilization of the polyplex in the blood circulation. In fact, we previously developed cyclodextrin (CD)/polyamidoamine dendrimer conjugates (CDE) as siRNA carriers, and the interaction of CDE/siRNA was decreased by the conjugation with folate-polyethylene glycol, leading to the destabilization. To overcome this dilemma, in this study, folate-appended polyrotaxanes (Fol-PRX) were developed. Fol-PRX stabilized CDE/siRNA polyplex by intermolecularly connecting CDE molecules through a host–guest interaction between adamantane at the terminals of Fol-PRX and β-CD in the polyplex. Moreover, the intermolecular connection of the polyplex with Fol-PRX provided movable folate moieties on the surface. As a result, Fol-PRXs enhanced the in vivo antitumor activity of the polyplex after intravenous administration, suggesting their utility as the dual-functional materials for systemic delivery of siRNA polyplexes.
Free cholesterol acts as an endogenous agonist for estrogen-related receptor α (ERRα), a nuclear receptor that regulates osteoclastogenesis. Because stimulation of macrophages with receptor activator of nuclear factor κB ligand (RANKL) induces an overload of free cholesterol and activates ERRα, we hypothesized that direct removal of cellular cholesterol would suppress osteoclastogenesis. In this study, the effect of 2-hydroxypropyl β-cyclodextrin (HP-β-CD), a highly water-soluble cyclic glucopyranose, and β-CD-threaded polyrotaxanes (PRXs), supramolecular polymers designed to release threaded β-CDs in acidic lysosomes, on RANKL-induced cholesterol overload and osteoclast differentiation of murine macrophage-like RAW264.7 cells were investigated. PRXs suppressed RANKL-induced cholesterol overload. Additionally, RANKL-induced osteoclast differentiation of RAW264.7 cells was inhibited by PRXs. In contrast, HP-β-CD did not reduce cholesterol levels or inhibit osteoclast differentiation in RAW264.7 cells. Gene expression analysis of osteoclast markers suggested that PRXs suppress only the early stage of osteoclast differentiation, as PRXs cannot be internalized into multinucleated osteoclasts. However, modification of PRXs with cell-penetrating peptides facilitated their cellular uptake into multinucleated osteoclasts and inhibited osteoclast maturation. Thus, PRXs are promising candidates for inhibiting osteoclast differentiation by suppressing cholesterol overload and may be useful for treating osteoporosis or other bone defects caused by the overactivity of osteoclasts.
Cas9 ribonucleoprotein (Cas9 RNP) is a promising genome editing tool, however its biological utility reuires the development of safe, efficient, and easy-to-use non-viral carriers. Cas9 RNP has a complicated conformation and charge distribution, resulting in low complexation with carriers. In addition, intracellular uptake, endosome escape, release, and nuclear translocation of Cas9 RNP are required. Here, we report the development of polyrotaxane-based supramolecular carriers, aminated polyrotaxanes (amino-PRXs), that efficiently form complexes with Cas9 RNP via their autonomous transforming properties (1st generation; 1 G). Further, the amino groups of amino-PRXs are optimized to provide endosome-escape ability (2 G) via transforming to highly cationic particles in the endosome. Moreover, intracellular degradation properties are provided for Cas9 RNP release (3G–5 G) resulting in released Cas9 RNP becoming localized in the nucleus. Finally, we demonstrate that this optimized amino-PRX (5 G) facilitates highly efficient genome editing both in vitro and in vivo with significant usability, suggesting that amino-PRX (5 G) is a promising platform for the development of non-viral Cas9 RNP carriers.
Synthetically trapping kinetically varied (super)structures of molecular assemblies and amplifying them to the macroscale is a promising, yet challenging, approach for the advancement of meta-stable materials. Here, we demonstrated a concerted kinetic trapping design to timely resolve a set of transient polypseudorotaxanes in solution and harness a crop of them via micro-crystallization. By installing stopper or speed bump moieties on the polymer axles, meta-stable polypseudorotaxanes with segmented cyclodextrin blocks were hierarchically amplified into crystalline networks of different crosslinking densities at mesoscale and viscoelastic hydrogels with 3D-printability in bulk. We demonstrated simultaneous 3D-printing of two polypseudorotaxane networks from one reactive ensemble and their conversion to heterogeneous polyrotaxane monoliths. Spatially programming the macroscale shapes of these heterogeneous polyrotaxanes enabled the construction of moisture-responsive actuators, in which the shape morphing originated from the different numbers of cyclodextrins interlocked in these polyrotaxane networks.
Cyclodextrin-based polymers (CDPs) are amphiphilic macromolecules owing to their unique structure and the presence of multiple hydroxyl groups. These polymers have become increasingly popular in applications ranging from biomedicine to environmental science. In this review, four types of CDPs are distinguished based on their structural characteristics: cyclodextrin-based polyrotaxanes, grafted cyclodextrin polymers, cross-linked cyclodextrin polymers, and cyclodextrin-originated star/multi-arm polymers. The discussion includes the syntheses and properties of several derivatives of CDPs. In addition to low toxicity, good bioavailability and biodegradability, and the feasibility of inclusion complexation and chemical modification of cyclodextrins, these polymers have prominent advantages, such as stimuli responsiveness and the ability to form well-defined aggregated nanostructures, which make them promising candidates for applications related to controlled and targeted drug delivery. In this review, we have highlighted the evolution, unique types, and properties of various CDPs, which are core aspects that differ from previously published reviews. Additionally, the synergistic effect of cyclodextrin and cross-linking monomers, as well as their use as drug carriers over the past decade, have been emphasized.
The crystal structure of γ-cyclodextrin complexes with several organic compounds have been investigated by X-ray powder method. A two-dimensional tetragonal unit cell having a=b=27.2 Å and a two-dimensional hexagonal unit cell having a=b=32.7 Å, were reasonably proposed for hydrated and anhydrous γ-dextrin complexes, respectively. The change of the crystal structure caused by dehydration seemed to be resulted from the change of the packing arrangement of circular cylinders that are made by coaxial alignment of the dextrin molecules. Results obtained were tentatively applied to consider the 81-helical configuration of amylose.
A pair of radial [4]catenane isomers interlocked with two CB[6]s and one β-CD is reported. Due to the different positions of the tightly bound CB[6]s, shuttling dynamics of the β-CD between the two biphenyl stations are different in the isomers.
Mechanically interlocked polymers (MIPs), such as polyrotaxanes and polycatenanes, are polymer architectures that incorporate a mechanical bond. In a polyrotaxane, the mechanical bond is the result of a linear dumbbell component threaded through a ring, while in a polycatenane, it is the consequence of interlocked ring components. The interlocked nature of these architectures can result in high degrees of conformational freedom and mobility of their components, which can give rise to unique property profiles. In recent years, the synthesis and studies of a range of MIPs has allowed researchers to build an initial understanding of how incorporating mechanical bonds within a polymer structure impacts its material properties. This Review focuses on the understanding of these structure–property relationships with an outlook towards their applications, specifically focusing on four main classes of MIPs: polyrotaxanes, slide-ring gels, daisy-chain polymers and polycatenanes. Incorporating the mechanical bond into polymer architectures allows access to polymers with high-mobility elements, leading to unique material properties. This Review outlines the structure–property relationships of materials based on either polyrotaxanes (including slide-ring materials and daisy-chain polymers) or polycatenanes, and looks towards future applications and technologies.
The sliding dynamics of one- or multi-rings along a semiflexible cyclic polymer in radial poly[n]catenanes is investigated using molecular dynamics simulations. The fixed and fluctuating (non-fixed) semiflexible central cyclic polymers are considered, respectively. Increasing the bending energy of central cyclic polymer, for the fixed case, the diffusion coefficient increases monotonically due to the reduction of the tortuous sliding path, while for the fluctuating case, the diffusion coefficient decreases. It indicates that the contribution of the polymer fluctuation is suppressed by a further increase in the stiffness of the central cyclic chain. Compared with one ring case, the mean-square displacement of the multi-rings exhibits a unique subdiffusive behavior at the intermediate time scales due to the repulsion between two neighboring rings. In addition, for the multi-rings system, the whole set of rings exhibit relatively slower diffusion, but faster local dynamic of threading rings and rotational diffusion of central cyclic polymer arise. These results may help us understand the diffusion motion of ring in radial poly[n]catenanes from a fundamental view and control the sliding dynamics in molecular designs.
We previously found that acid-labile polyrotaxane containing methylated β-cyclodextrin (Me-PRX) induces endoplasmic reticulum (ER) stress-related autophagy and autophagic cell death. Me-PRX-induced autophagic cell death occurs even in apoptosis-resistant cells; tumor-targeted Me-PRX delivery could thus be an effective cancer treatment approach. In this study, antibody–supermolecule conjugates, consisting of a tumor-specific antibody and Me-PRX, were designed to achieve a tumor-specific delivery of Me-PRX. Trastuzumab, a monoclonal antibody against HER2 expressed in various malignant tumors, was selected as a tumor-targeting antibody, and phenyl maleimide group-modified Me-PRX (Mal-Me-PRX) was conjugated to the cysteine residue of the reduced Trastuzumab to obtain a Trastuzumab–Me-PRX conjugate (Tras-Me-PRX). The cellular association of Tras-Me-PRX to HER2-expressing tumor cells was remarkably greater than that of unmodified Me-PRX. Moreover, Tras-Me-PRX effectively reduced the viability of HER2-expressing tumor cells at a lower concentration compared to the unmodified Me-PRX. In conclusion, antibody–Me-PRX conjugates are regarded as a new class of antibody–drug conjugates that would contribute to the chemotherapy of cancers.
Acetylated α-cyclodextrin (α-CD)/poly(ethylene glycol) (PEG)-based polyrotaxanes (Ac-PRXs) with varying degrees of acetylation (DA) and molecular weight of axle PEG were synthesized and their solubility in aqueous solutions was investigated. Ac-PRXs with low DA (less than 35%) were dissolved in aqueous solutions without considering the molecular weight of axle PEG, whereas Ac-PRXs with high DA (more than 40%) and low molecular weight of axle PEG (less than 35000) were precipitated into the solutions. Interestingly, Ac-PRXs with high DA and high molecular weight of axle PEG (100000) exhibited a colloidal dispersion in aqueous solutions. It is considered that the threaded acetylated α-CDs formed hydrophobic microenvironments via hydrophobic interactions and the noncovered segments of axle PEGs provided colloidal stability. Furthermore, the potential application of Ac-PRX100k as a drug carrier was examined and it was established that Ac-PRX100k can encapsulate a hydrophobic drug. Accordingly, acetylation of PRXs is a viable approach to promote solubility in aqueous solutions and prepare self-assembled nanoparticles.
Failure of autophagy induction results in the accumulation of abnormal mitochondria to cause neurodegenerative diseases. Artificial autophagy activation by the mitochondrial delivery of polyrotaxane with autophagy inducing activity is achived...
Nanosheet materials have recently attracted the attention of researchers because of their unique physical properties. In the present study, we have reported a novel methodology to fabricate isolated nanosheet materials, called pseudo-polyrotaxane (PPR) nanosheets, which are formed by biocompatible complexation between α-cyclodextrin (CD) and poly(ethylene glycol) (PEG) for the first time. When the molecular weight of axis PEG was changed in the range between 2k and 6k, isolated PPR nanosheets were obtained with thicknesses ranging from 14.6 to 33.8 nm, depending on the axis PEG length. We found that uncovering the axis ends led to the formation and isolation of PPR nanosheets when the binding constant between the axis ends and α-CD was changed. Additionally, the PPR structures consisting of multiarm PEG indicated that the uncovered parts of the axis near the branched point also promoted the formation of isolated PPR nanosheets. Uncovering the parts of the axis polymer is essential for fabricating the isolated PPR nanosheets because the uncovered parts of the axis suppress the crystal growth of α-CD in the axis direction and results in the isolation of PPR nanosheets.
Charged catenane has attracted an ever increasing attention as prototypical molecular switches in their synthesis and function. In this study, β-cyclodextrin and ionic liquid interlocking mechanically with each other are carried out to form charged catenane through ionic hydrogen bonding. When the charged catenane is treated with Cu(II) ion, it will generate blue precipitate quickly with a rigid and robust structure. Given the synthesized molecule bears good electroactivity and stability, it is used as modifies on the surface of the bare glassy carbon electrode and further performed as an electrochemical probe. The probe shows a clear chiral discrepancy in the response of oxidation peak current (IP) toward four isomers, including tryptophan, tyrosine, cysteine, and malic acid. That is, L form has a much higher IP and meantime it is hard to observe the electrochemical signal for D form. More interestingly, the recognition ability between L and D forms of tryptophan can be reversed in the buffer solution with different pH values. These results show that the dynamic switch process of steric hindrance based on the catenane can enlarge the chiral discrepancy. In a word, we believe that this study would enrich the synthetic pathways of electroactive molecules and lead to a deeper fundamental understanding of their function.
The molecular mobility of threading cyclic molecules (e.g. α-cyclodextrins) along a linear polymer chain (e.g. poly(ethylene glycol)) in polyrotaxanes is a unique feature for biomaterials with dynamic functionality. Surfaces with molecular mobility can be obtained by introducing polyrotaxanes. The molecular mobility of polyrotaxane-based surfaces can be modulated by changing the number of threading cyclic molecules and modifying their functional groups. Biological ligands modified with α-cyclodextrins exhibit increased multivalent interactions with their receptors due to the molecular mobility of the latter. Furthermore, polyrotaxane-based surfaces not only improve the initial response of cells via multivalent interactions but also affect cytoskeleton formation and the inherent quality of cells, including differentiation. Such polyrotaxane surfaces can emerge as new biointerfaces that can adapt to the dynamic biological nature.
Autophagy plays a pivotal role in the development and prevention of numerous diseases, and the induction of autophagy is regarded as a potential therapeutic approach for intractable diseases. Methylated β-cyclodextrins (Me-β-CDs) are known to destabilize the plasma membrane through the removal of cholesterols via inclusion complexation, which leads to the induction of apoptosis. In this study, the induction of autophagy by Me-β-CDs-threaded acid-labile polyrotaxane (Me-PRX) that can release the threaded Me-β-CDs in response to acidic pH in lysosomes was investigated. We hypothesized that the Me-β-CDs released from the Me-PRX interact with the membrane of organelles and cause autophagy. The Me-PRX preferentially accumulated in endoplasmic reticulum (ER) and caused ER stress, which was confirmed by gene expression analysis and the expression of an ER stress-marker protein. Accompanying the ER stress, cells treated with Me-PRX showed autophagy, which was not observed in cells treated with non-labile Me-PRX, other chemically modified PRXs, or free Me-β-CD. Furthermore, the Me-PRX treatment induced autophagic cell death and caused cell death even in apoptosis-resistant cells. Overall, this study demonstrates that the acid-labile Me-PRX induces ER stress-mediated autophagic cell death, and the Me-PRX would be a promising candidate to induce effective cell death in apoptosis-resistant malignant tumors.
Niemann-Pick type C (NPC) disease is a fatal metabolic disorder characterized by the lysosomal accumulation of cholesterol. Although 2-hydroxypropyl β-cyclodextrin (HP-β-CD) promotes the excretion of cholesterol and prolongs the life span in animal models of NPC disease, it requires extremely high dose. We developed acid-labile β-CD-based polyrotaxanes (PRXs) comprising multiple β-CDs threaded along a polymer chain capped with acid-cleavable stopper molecules for potentiating therapeutic efficacy of β-CD in NPC disease. The acid-labile PRXs dissociate under the acidic lysosomes and release threaded β-CDs in lysosomes, which promotes cholesterol excretion in NPC disease model cells at lower concentration than HP-β-CD. In this study, the therapeutic effect of the PRXs in a mouse model of NPC disease was investigated. Weekly administration of the PRXs significantly prolonged the life span and suppressed neurodegeneration in mice, even at a dose of 500mg/kg, a markedly lower dose than previously reported for HP-β-CD. Detailed analysis of tissue cholesterol revealed that PRX treatment markedly suppressed the tissue accumulation of cholesterol in the NPC mouse model, but did not alter cholesterol content in wild-type mice. Acid-labile PRX is therefore a promising candidate for potentiating the efficacy of β-CD in the treatment of NPC disease.
In the retinal pigment epithelium of patients with age-related macular degeneration (AMD), excess N-retinylidene-N-retinylethanolamine (A2E), a dimer of all-trans-retinal, accumulats to induce inflammatory cytokine secretion and phototoxic effects. Therefore, the reduction of intracellular A2E is a promising approach for the prevention and treatment of AMD. In this study, acid-labile β-cyclodextrin (β-CD)-threaded polyrotaxanes (PRXs) were synthesized and investigated their effects on the removal of A2E accumulated in retinal pigment epithelium cells (ARPE-19) in comparison to nonlabile PRXs and 2-hydroxypropyl β-CD (HP-β-CD) were examined. GC-MS and HPLC studies strongly suggest that the acid-labile PRXs dissociated into their constituent molecules in cells by lysosomal acidification and threaded β-CDs were considered to be released from the PRXs. The released β-CDs formed an inclusion complex with A2E, which promoted the excretion of A2E. Indeed, the acid-labile PRXs effectively reduced intracellular A2E level at approximately a 10-fold lower concentration than HP-β-CD. Accompanied with A2E removal, the toxicity and phototoxicity of A2E were attenuated by treatment with acid-labile PRXs. Because the nonlabile PRX failed to reduce intracellular A2E level and attenuate phototoxicity, intracellular release of threaded β-CDs from the acid-labile PRX might contribute to reducing intracellular A2E. We conclude that acid-labile PRXs are promising candidates for the treatment of macular diseases through the removal of toxic metabolites.
We developed stimuli-labile polyrotaxanes (PRXs) composed of β-cyclodextrin (β-CD), Pluronic as an axle polymer, and acid-cleavable N-triphenylmethyl groups as bulky stopper molecules, and found that the PRXs are potent therapeutics for Niemann-Pick type C disease, because the PRX can effectively reduce intracellular cholesterol through the intracellular release of threaded β-CDs. In general, the PRXs need to be chemically modified with hydrophilic functional groups because PRXs are not soluble in aqueous media. Herein, four series of oligo(ethylene glycol)s (OEGs) with different ethylene glycol repeating unit (2 or 3) and chemical structure of OEG terminal (hydroxy or methoxy) were modified onto the threaded β-CDs in PRX. The effects of the structure of OEG on the aqueous solubility, toxicity, and cellular internalization efficiency of OEG-modified PRXs were investigated to optimize the chemical structure of OEG. The hydroxy-terminated OEG-modified PRXs showed excellent solubility in aqueous media and no toxicity, regardless of the number of ethylene glycol repeating units. In the case of the methoxy-terminated OEG-modified PRXs, sufficient solubility in aqueous media and negligible toxicity were observed when the number of ethylene glycol repeating units was 3, while low solubility and toxicity were observed when the ethylene glycol repeating unit was 2. Additionally, cellular uptake levels of methoxy-terminated OEG-modified PRXs in RAW264.7 cells were higher than those of hydroxy-terminated OEG-modified PRXs. Consequently, the chemical structure of the OEG strongly affects the chemical and biological properties of the PRXs, and that a methoxy-terminated OEG with 3 ethylene glycol repeating units is the most preferable modification of PRXs, since the resultant PRX is sufficiently soluble in aqueous media, non-toxic, and possesses high cellular internalization efficiency.
Polyrotaxanes (PRXs) composed of threading methylated cyclodextrins exhibit a temperature-dependent reversible phase transition across the lower critical solution temperature (LCST) in water. In this study, a variety of methylated β-cyclodextrin (β-CD)-threaded PRXs (Me-PRXs) with different number of methyl groups modified on the β-CD and with different compositions of axle polymers (i.e., Pluronic, PEG-b-PPG-b-PEG) were synthesized, and the relationship between the LCST value and structural parameters was investigated. The LCST values of the Me-PRXs decreased with an increasing the methylation degree of the threading β-CD. Differential scanning calorimetric measurements demonstrated that the temperature-induced phase transition of the Me-PRXs occurred due to the dehydration of the threading methylated β-CD moieties and the subsequent hydrophobic interactions. Interestingly, the Me-PRXs formed coacervate droplets above their LCST. Altogether, we concluded that the LCST values of the Me-PRXs varied according to their structural parameters and could be applied as temperature-responsive biomaterials or therapeutics.
In 2008, we reported a new class of pillar-shaped macrocyclic hosts, known as "pillar[n]arenes". Today, pillar[n]arenes are recognized as key players in supramolecular chemistry because of their facile synthesis, unique pillar shape, versatile functionality, interesting host-guest properties, and original supramolecular assembly characteristics, which have resulted in numerous electrochemical and biomedical material applications. In this Review, we have provided historical background to macrocyclic chemistry, followed by a detailed discussion of the fundamental properties of pillar[n]arenes, including their synthesis, structure, and host-guest properties. Furthermore, we have discussed the applications of pillar[n]arenes to materials science, as well as their applications in supramolecular chemistry, in terms of their fundamental properties. Finally, we have described the future perspectives of pillar[n]arene chemistry. We hope that this Review will provide a useful reference for researchers working in the field and inspire discoveries concerning pillar[n]arene chemistry.
We demonstrated a new strategy for efficient preparation of polypseudorotaxanes (PpRXs) and polyrotaxanes (PRXs) with cyclodextrin derivatives, 2,6-di-O-methyl-cyclodextrins (DM-CyDs), by utilizing the cloud points of DM-CyDs. DM-α-CyD and DM-β-CyD formed PpRXs with polyethylene glycol (PEG) and polypropylene glycol (PPG) in water at >50 °C and >35 °C, respectively, but did not at room temperature. Meanwhile, randomly methylated β-CyD (RM-β-CyD) and 2,3,6-tri-O-methyl-β-CyD (TM-β-CyD) did not form PpRX with PPG at higher temperature. The driving force of thermoresponsive formation of DM-CyD PpRXs was derived from hydrophobic interaction of methyl groups and a hydrogen bond of hydroxyl groups formed by adjacent DM-CyD molecules. Furthermore, in one pot, DM-CyD PRXs were synthesized by capping the PpRXs with bulky ends in high yields.
A UV-cleavable supramolecular cross-linker was designed to effectively control the mechanical strength of photocurable resin plastics. The resin monomer-soluble polyrotaxane (PRX) cross-linker was synthesized by introducing a hydrophobic n-butyl group and a cross-linkable methacrylate group in α-cyclodextrin threading to a polyethylene glycol containing UV-cleavable end groups. The UV-cleavable PRX cross-linker was completely dissolved in 2-hydroxyethytl methacrylate (HEMA) and camphorquinone, representative photocurable resin components. The dumbbell-like stiff resin plastic was prepared by irradiating the mixture with 450 nm blue light. The stiffened resin plastic maintained its ultimate tensile strength (UTS) under visible light irradiation. However, the UTS of the resin plastic was remarkably decreased to 40% of the original value once the plastic was exposed to 254 nm UV light. This indicates that the suggested UV-cleavable PRX cross-linker is effective in modulating the mechanical strength of photocurable resin plastics.
Cyclodextrins (CyDs) include linear polymers such as polyethylene glycol (PEG), polypropylene glycol (PPG) and polyethyleneimine (PEI) and spontaneously form supramolecular inclusion complexes, namely polypseudorotaxanes. When both ends of the polymer chains in polypseudorotaxanes are covalently capped with bulky molecules, CyDs are trapped in and cannot be dethreaded from the assembly, giving the so-called polyrotaxane. Recently, a large number of drug carriers based on polyrotaxanes and polypseudorotaxanes have been developed for various drugs, i.e., low molecular weight drugs, protein drugs, gene drugs and nucleic acid drugs. In this review, we introduce various applications of polyrotaxanes and polypseudorotaxanes as drug delivery techniques such as drug absorption, controlled release and drug targeting for various drugs.
pH-sensitive polymeric materials are rational design for therapeutics, because they can change their intrinsic properties selectively in response to pH changes in targeted tissues and organelles, such as tumor tissue, skin, inflammation site, and intracellular endosomes and lysosomes. Herein, we described a novel acid-labile Pluronic/β-cyclodextrin (β-CD)-based polyrotaxane (PRX) showing the dissociation of supramolecular interlocked structure in response to a pH change with a narrow acidic range for biomedical applications. Acid cleavable ketal linkages were introduced into both terminals of the axle polymer of the PRX (ket-PRX) to acquire acid sensitivity. The ket-PRX was sufficiently stable and maintain its supramolecular structure at physiologic condition (pH 7.4), whereas dissociation of the ket-PRX was observed at weakly acidic pH condition (pH 5.0) due to the cleavage of the ketal linkages. Concomitant with this supramolecular dissociation it was found that the ket-PRX released β-CDs to form an inclusion complex with guest molecules specifically at weakly acidic pH conditions. As one of the possible intracellular therapeutic applications, it was confirmed that the ket-PRX showed superior reduction of lysosomal cholesterols in Niemann-Pick type C disease in comparison to β-CD derivatives, presumably due to the local release of β-CDs from the ket-PRX in response to weakly acidic pH in endosomes and/or lysosomes. Altogether with these results, it is concluded that the ket-PRX exerting pH-sensitive dissociation is an attractive candidate as potential therapeutic biomaterials.
Polyrotaxanes (PRXs) are a class of supramolecular threaded macromolecules, in which cyclic molecules are threaded onto the main- or side-chain of polymers. To date, various studies have been conducted on the synthesis of PRXs, and various combinations of cyclic molecules and polymers that can form a PRX have been discovered. Among these combinations, PRXs composed of cyclodextrins (CDs) and a linear polymer have attracted much attention and have been investigated by many researchers. Because of the non-covalently associated characteristic of PRXs, these supermolecules exhibit unique properties, such as the dynamic motion of the threaded cyclic molecules along a polymer axle and complete dissociation of the supramolecular structure, that are never observed in other synthetic polymers. These inherent properties of PRXs are of interest in the design of novel biomaterials, such as hydrogels, scaffolds in tissue engineering, drug delivery carriers, and polymer-drug conjugates. Thus, various studies have been conducted to utilize PRXs as a framework for biomaterials. In this review, we describe the recent progress in biomaterial application of PRXs such as drug delivery and gene delivery.
Pillar[n]arenes, which we first synthesized and named in 2008, are new pillar-shaped macrocyclic hosts. Pillar[n]arene homologues with n = 5-10 have already been synthesized, but the cyclic pentamers, i.e., pillar[5]arenes, and cyclic hexamers, i.e., pillar[6]arenes, have been most widely used because these can be obtained in good yields. To date, nearly all pillar[n]arene-based supramolecular assemblies have been constructed using pillar[5]- and pillar[6]arene scaffolds. In this feature article, we describe supramolecular assemblies built using host-guest interactions depending on the cavity sizes of pillar[5]- and pillar[6]arenes. We first discuss the effects of the type of substituents on the rims of pillar[5]- and pillar[6]arenes on their solubilities, functionalities and host-guest properties. We then discuss supramolecular assemblies based on their host-guest properties and pillar-shaped architectures.
Poly(ethylene glycol)s (PEG) of high molecular weights were found to form complexes with α-cyclodextrin (α-CD) in aqueous solutions to give gels in a wide range of concentrations. The time of gelation decreases with increase in α-CD and PEG concentrations, indicating that the gels formed during complex formation between α-CD and PEG chains. The time of gelation increases with increase in the molecular weight of PEG, indicating that the PEG chains penetrate α-CD cavities from the ends of PEG and are included by α-CDs. X-Ray powder diffraction studies showed that the gel consists of both complexed α-CD and uncomplexed α-CD, indicating partial inclusion of PEG chains by α-CD. The configuration of the gels and mechanism for the gelation are discussed, and it is suggested that α-CD-PEG complexes act as physical cross-links during gelation. The gel-melting temperature increases with increase in PEG molecular weights and α-CD concentrations, and decreases with increase in PEG concentrations, suggesting that gelation results from the formation of longer or shorter domains of α-CD–PEG inclusion complexes, respectively.
For years, great efforts have been made by scientists to construct various novel interlocked supramolecular systems. A wide range of pseudorotaxanes and rotaxanes [1-7], full of challenging constructions and potential applications in areas such as nanostructured functional materials [8,9], molecular switches [10-14], molecular logic gates [15-19], molecular wires [20-22], memory devices Willner [23] and biomedical applications [24,25] have been reported recently. Cyclodextrin polyrotaxantes or polypseudorotaxanes there are not only nice and interesting supramolecular architectures but they also have a high interest for biomaterials application that allow to opened up new approaches for tissue regeneration, drug and gene delivery. The use of the supramolecular cyclodextrins complexes has given rise to interesting studies to design and develop new biomaterials with advanced properties. In this review we will update the recent advances in the use of CDs-supramolecular structures for develop new and advanced drug and gene delivery systems and for use in tissue engineering.
Five polyrotaxanes were synthesized by threading 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) onto a variety of α,ω-ditriethylenediamino-N-carbamoyl-poly-(ethylene oxide)-block-poly(propylene oxide)-block-poly(ethylene oxide) (Pluronic) triblock copolymers using a two-pot strategy under heterogeneous, nonaqueous conditions. The threaded HP-β-CD units were retained on the pseudopolyrotaxane precursors by end-capping the branched diamine termini with sodium 2,4,6-trinitrobenzene sulfonate. Inclusion of the Pluronic copolymers within the HP-β-CD cavities was more favorable in nonpolar solvents, such as diethyl ether and n-hexane, both of which gave better coverage ratios than polar solvents. (1)H NMR and MALDI-TOF were used to estimate the average molecular weights of the purified polyrotaxane products. A globular morphology of aggregated polyrotaxanes was observed by tapping-mode AFM imaging of dried samples. Treatment of Niemann-Pick C (NPC) type 2-deficient fibroblasts with the polyrotaxane derivatives produced substantial reductions in sterol accumulation, as seen by diminished filipin staining in these cells, suggesting that Pluronic-based polyrotaxanes may be promising vehicles for delivery of HP-β-CD to cells with abnormal cholesterol accumulation.