Hydrogen-Bonded Structure of Water in the Loop of Anchored Polyrotaxane Chain Controlled by Anchoring Density.

Hydrogen-bonded network of water surrounding polymers is expected to be one of the most relevant factors affecting biocompatibility, while the specific hydrogen-bonded structure of water responsible for biocompatibility is still under debate. Here we study the hydrogen-bonded structure of water in a loop-shaped poly(ethylene glycol) chain in a polyrotaxane using synchrotron soft X-ray emission spectroscopy. By changing the density of anchoring molecules, hydrogen-bonded structure of water confined in the poly(ethylene glycol) loop was identified. The XES profile of the confined water indicates the absence of the low energy lone-pair peak, probably because the limited space of the polymer loop entropically inhibits the formation of tetrahedrally coordinated water. The volume of the confined water can be changed by the anchoring density, which implies the ability to control the biocompatibility of loop-shaped polymers.

Application of DNA-Alkylating Pyrrole-Imidazole Polyamides for Cancer Treatment.

Pyrrole-imidazole (PI) polyamides, which target specific DNA sequences, have been studied as a class of DNA minor-groove-binding molecules. To investigate the potential of compounds for cancer treatment, PI polyamides were conjugated with DNA-alkylating agents, such as seco-CBI and chlorambucil. DNA-alkylating PI polyamides have attracted attention because of their sequence-specific alkylating activities, which contribute to reducing the severe side effects of current DNA-damaging drugs. Many of these types of conjugates have been developed as new candidates for anticancer drugs. Herein, we review recent progress into research on DNA-alkylating PI polyamides and their sequence-specific action on targets associated with cancer development.

Molecular Recognition of Fluorescent Probe Molecules with a Pseudopolyrotaxane Nanosheet.

Pseudopolyrotaxane nanosheets (PPRNS) are ultrathin two-dimensional (2D) materials fabricated via supramolecular self-assembly of ß-cyclodextrin (ß-CD) and poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) triblock copolymers. In this study, the molecular loading of various fluorescent probe molecules onto PPRNS was systematically investigated. 1H NMR study for R6G absorption to PPRNS indicated that the small hydrophobic groups, such as the methyl group, of R6G were absorbed by PPRNS. Consistently, the fluorescent probes without methyl groups were not absorbed. These results indicate that PPRNS has a molecular recognition absorption property based on the host-guest interaction of the functional groups on probe molecules and molecular-sized spaces of PPRNS surfaces, which may be vacant ß-CDs and voids between ß-CD columns. The absorbed amount of the molecular probes onto PPRNS was investigated by UV-vis spectra, and the absorption behavior could be described well by the Langmuir absorption isotherm. This is consistent with the suggested model that the probes are absorbed onto the PPRNS surfaces. This study demonstrates that PPRNSs can be applied as adsorbents for toxic compounds, drug delivery systems, and 2D sensors.

Precise control of cyclodextrin-based pseudo-polyrotaxane lamellar structure via axis polymer composition.

Self-assembly of cyclodextrin (CD) with guest polymers has attracted much attention owing to its biocompatibility and accessibility. In this study, we investigate the composition effect of poly(ethylene oxide)m-b-poly(propylene oxide)n-b-poly(ethylene oxide)m (EOmPOnEOm) triblock copolymers on lamellar or plate structures formed by complexation with ß-CD. EO5PO29EO5, EO14PO29EO14, and EO75PO29EO75 show periodic lamellar morphology consisting of single-crystalline pseudo-polyrotaxane (PPR) nanosheets with a thickness equal to the central PO length. This is because ß-CDs selectively cover the PO component and cause the microphase separation between ß-CD and EO layers. The thickness of the EO layers increases linearly with increasing number of EO units, which suggests that the EO chains are constrained into virtual cylinders with the diameter of the ß-CD. This means that we can precisely control the thickness of both the crystal (ß-CD and PO) and the amorphous (EO) layers in the lamellar structure. In contrast, EO2PO29EO2 forms a thin plate structure, where not only PO but also EO chains are covered with ß-CD. Furthermore, the length of the central PO component is necessary to form the lamellar structure with the phase separation between the ß-CD and EO layers. These findings provide a more fundamental understanding to enhance the variety and applicability of CD-based self-assembled materials.

Photoprotective Effects of Selected Amino Acids on Naproxen Photodegradation in Aqueous Media.

It is important to develop a photostabilization strategy to ensure the quality of photosensitive compounds, including pharmaceuticals. This study focused on the protective effects of 20 amino acids on the photodegradation of naproxen (NX), a photosensitive pharmaceutical, to clarify the important nature of a good photostabilizer. Our previous report indicated the photodegradability of NX and the protective effects of some antioxidants on its photodegradation, therefore, this compound was used as a model compound. The degradation of NX in aqueous media during ultraviolet light (UV) irradiation and the protective effects of selected amino acids were monitored through high-performance liquid chromatography (HPLC), equipped with a reverse-phase column. Addition of cysteine, tryptophan, and tyrosine induced the significant suppression of NX photodegradation after UV irradiation for 3 h (residual amount of NX; 15.35%, 6.82%, and 15.64%, respectively). Evaluation of the antioxidative activity and UV absorption spectrum showed that cysteine suppressed NX degradation through its antioxidative ability, while tryptophan and tyrosine suppressed it through their UV filtering ability. Furthermore, three amino acids at higher concentrations (more than 100 µmol/L) showed more protective effects on NX photodegradation. For 10 mmol/L, residual amounts of NX with cysteine, tryptophan, and tyrosine were 58.51%, 69.34%, and 82.40%, respectively. These results showed the importance of both photoprotective potencies (antioxidative potency and UV filtering potency) and stability to UV irradiation for a good photostabilizer of photosensitive pharmaceuticals.

DNA Alkylation of the RUNX-Binding Sequence by CBI-PI Polyamide Conjugates*.

Many types of molecular targeted drugs that inhibit cancer growth by acting on specific molecules have been developed. The runt-related transcription factor (RUNX) family, which induces cancer development by binding to a specific DNA sequence, has attracted attention as a new target for cancer treatment. We have developed Chb-M', which targets the RUNX-binding sequence. Chb-M' was developed by conjugating pyrrole-imidazole (PI) polyamides and chlorambucil as an anticancer agent. It was recently reported that Chb-M' had a remarkable anticancer effect in vivo. In this study, to explore the possibility of an alternative structure, we designed a new series of CBI-PI polyamides, in which seco-CBI was applied as a DNA-alkylating agent. We examined the characteristics of the CBI-PI polyamides targeting the RUNX-binding sequence and found that these conjugates have great potential for cancer treatment.

Synthesis of Poly(Methyl Methacrylate)-Based Polyrotaxane via Reversible Addition-Fragmentation Chain Transfer Polymerization.

A polyrotaxane (PR) with poly(methyl methacrylate) (PMMA) as the main chain polymer was prepared via reversible addition-fragmentation chain transfer (RAFT) polymerization. Because of the special mechanism of RAFT, the suprastructure of a PMMA-based PR is established by synthesizing inclusion complexes of methyl methacrylate and gamma-cyclodextrin (γCD) into the middle of the poly-N-(3-dimethylamino) propyl methacrylamide segments. The presence of threaded γCD was determined via diffusion ordered spectroscopy from the alignment of the mobility of γCD and the main chain polymer. A PMMA-based PR with 2-20% CD coverage and a molecular weight of 7K-60K g/mol of PMMA-based PR was synthesized with a targeted molecular structure by mediating the RAFT polymerization. The PMMA-based PR prepared in this study is expected to be suitable for wide applications of tough materials with good heat resistance. Moreover, the investigation of this synthetical approach opened possibilities for more variety of PR with controllable properties.

Autonomously isolated pseudo-polyrotaxane nanosheets fabricated via hierarchically ordered supramolecular self-assembly.

We succeeded in obtaining autonomously isolated nanosheets consisting of pseudo-polyrotaxane (PPR) fabricated via hierarchically ordered supramolecular self-assembly of ß-cyclodextrin and a poloxamer by introducing charged groups to the axis ends of the poloxamer. The isolated PPR nanosheets exhibited a tunable structural coloration and were aligned using a strong magnetic field.

Molecular Characteristics of DNA-Alkylating PI Polyamides Targeting RUNX Transcription Factors.

The runt-related transcription factor (RUNX) family has been associated with cancer development. The binding of RUNX family members to specific DNA sequences is hypothesized to promote the expression of downstream genes and cause cancer proliferation. On the basis of this proposed mechanism of cancer growth, we developed conjugate 1, which inhibits the binding of RUNX to its target DNA. Conjugate 1 is a DNA-alkylating pyrrole-imidazole (PI) polyamide conjugate containing chlorambucil as an anticancer agent. Conjugate 1 was reported to have a marked anticancer effect in mouse models of acute myeloid leukemia. Although the effectiveness of 1 has been demonstrated in vivo, the detailed mechanism by which it alkylates DNA is unknown. Here, we chemically elucidated the molecular characteristics of conjugate 1 to confirm its potential as a RUNX-inhibiting drug. We also generated an alternative conjugate 2, which targets the same DNA sequence, by replacing one pyrrole with ß-alanine. Comparison of the characteristics of conjugates 1 and 2 suggested that reaction selectivity and binding affinity to the RUNX-binding sequence were improved by the introduction of ß-alanine. These findings indicate the possibility of DNA-alkylating PI polyamides as candidates for cancer chemotherapeutics.

Drastic Change of Mechanical Properties of Polyrotaxane Bulk: ABA-BAB Sequence Change Depending on Ring Position.

Polyrotaxane (PR), consisting of many ring molecules and an axis polymer, is a typical supramolecular structure with unique topological characteristics. In this study, we demonstrated the drastic change of the macroscopic mechanical properties depending on the ring position of PR in bulk. Poly(ethylene oxide)-b-poly(propylene oxide)-b-poly(ethylene oxide) (PEO-PPO-PEO) triblock copolymer was employed as an axis polymer to control the position of ß-cyclodextrin (ß-CD). To transfer the ß-CD positions, hydroxypropyl groups (HPPR) and hydrophobic trimethyl silyl groups (TMS-HPPR), which have hydrophilic and hydrophobic ß-CD, respectively, were synthesized. ß-CDs in HPPR were localized on a central PPO segment and formed crystal domains. The axis polymer of HPPR could not bridge ß-CD crystal domains, resulting in a melt state at high temperature. On the other hand, ß-CDs in TMS-HPPR were transferred to both PEO segments and formed crystal domains. The axis polymer in TMS-HPPR could bridge the ß-CD crystal domains, resulting in an elastic state even at high temperature. We succeeded in demonstrating the potential ability of PR: the macroscopic mechanical properties of PR can be changed from a melt state to an elastic one by manipulating the ring positions.

Evaluation of alkylating pyrrole-imidazole polyamide conjugates by a novel method for high-throughput sequencer.

N-Methylpyrrole-N-methylimidazole (PI) polyamides are a class of DNA minor groove binders with DNA sequence-specificity. DNA-alkylating PI polyamide conjugates are attractive candidates as anticancer drugs acting through DNA damage and its subsequent inhibition of cell proliferation. One example is a chlorambucil-PI polyamide conjugate targeting the runt-related transcription factor (RUNX) family. RUNX1 has pro-oncogenic properties in acute myeloid leukemia, and recently the chlorambucil-PI polyamide conjugate was demonstrated to have anticancer effects. Herein, we apply another DNA-alkylating agent, seco-CBI, to target the consensus sequence of the RUNX family. Two types of CBI conjugates were prepared and their binding properties were characterized by Bind-n-Seq analysis using a high-throughput sequencer. The sequencing data were analyzed by two methods, MERMADE and our new MR (motif identification with a reference sequence), and the resultant binding motif logos were as predicted from the pairing rules proposed by Dervan et al. This is the first report to employ the MR method on alkylating PI polyamide conjugates. Moreover, cytotoxicity of conjugates 3 and 4 against a human non-small cell lung cancer, A549, were examined to show promising IC50s of 120 nm and 63 nm, respectively. These findings suggest seco-CBI-PI polyamide conjugates are candidates for oncological therapy.

Genetic regulation of the RUNX transcription factor family has antitumor effects.

Runt-related transcription factor 1 (RUNX1) is generally considered to function as a tumor suppressor in the development of leukemia, but a growing body of evidence suggests that it has pro-oncogenic properties in acute myeloid leukemia (AML). Here we have demonstrated that the antileukemic effect mediated by RUNX1 depletion is highly dependent on a functional p53-mediated cell death pathway. Increased expression of other RUNX family members, including RUNX2 and RUNX3, compensated for the antitumor effect elicited by RUNX1 silencing, and simultaneous attenuation of all RUNX family members as a cluster led to a much stronger antitumor effect relative to suppression of individual RUNX members. Switching off the RUNX cluster using alkylating agent-conjugated pyrrole-imidazole (PI) polyamides, which were designed to specifically bind to consensus RUNX-binding sequences, was highly effective against AML cells and against several poor-prognosis solid tumors in a xenograft mouse model of AML without notable adverse events. Taken together, these results identify a crucial role for the RUNX cluster in the maintenance and progression of cancer cells and suggest that modulation of the RUNX cluster using the PI polyamide gene-switch technology is a potential strategy to control malignancies.

Perpendicular Orientation Control without Interfacial Treatment of RAFT-Synthesized High-χ Block Copolymer Thin Films with Sub-10 nm Features Prepared via Thermal Annealing.

In this study, a series of perpendicular lamellae-forming poly(polyhedral oligomeric silsesquioxane methacrylate-block-2,2,2-trifluoroethyl methacrylate)s (PMAPOSS-b-PTFEMAs) was developed based on the bottom-up concept of creating a simple yet effective material by tailoring the chemical properties and molecular composition of the material. The use of silicon (Si)-containing hybrid high-χ block copolymers (BCPs) provides easy access to sub-10 nm feature sizes. However, as the surface free energies (SFEs) of Si-containing polymers are typically vastly lower than organic polymers, this tends to result in the selective segregation of the inorganic block onto the air interface and increased difficulty in controlling the BCP orientation in thin films. Therefore, by balancing the SFEs between the organic and inorganic blocks through the use of poly(2,2,2-trifluoroethyl methacrylate) (PTFEMA) on the organic block, a polymer with an SFE similar to Si-containing polymers, orientation control of the BCP domains in thin films becomes much simpler. Herein, perpendicularly oriented BCP thin films with a χeff value of 0.45 were fabricated using simple spin-coating and thermal annealing processes under ambient conditions. The thin films displayed a minimum domain size of L0 = 11 nm, as observed via atomic force microscopy (AFM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Furthermore, directed self-assembly (DSA) of the BCP on a topographically prepatterned substrate using the grapho-epitaxy method was used to successfully obtain perpendicularly oriented lamellae with a half pitch size of ca. 8 nm.

Perpendicularly oriented sub-10-nm block copolymer lamellae by atmospheric thermal annealing for one minute.

The directed self-assembly (DSA) of block co-polymers (BCPs) can realize next-generation lithography for semiconductors and a variety of soft materials. It is imperative to simultaneously achieve many requirements such as a high resolution, orientation control of micro-domains, etch selectivity, rapid and mild annealing, a low cost, and compatibility with manufacturing for developing suitable BCPs. Here, we describe a new design for modified polysiloxane-based BCPs targeted for sub-10-nm-wide lines, which are able to form perpendicularly oriented lamellar structures in thin films. The hydroxyl groups in the side chains introduced in the polysiloxane block provide a good balance with the polystyrene surface free energy, thereby leading to the perpendicular orientation. Moreover, this orientation can be completed in only one minute at 130 °C in an air atmosphere. Oxygen plasma etching for the thin films results in the achievement of a line width of 8.5 nm.

Bacterial transport of colloids in liquid crystalline environments.

We describe the controlled transport and delivery of non-motile eukaryotic cells and polymer microparticles by swimming bacteria suspended in nematic liquid crystals. The bacteria push reversibly attached cargo in a stable, unidirectional path (or along a complex patterned director field) over exceptionally long distances. Numerical simulations and analytical predictions for swimming speeds provide a mechanistic insight into the hydrodynamics of the system. This study lays the foundation for using cargo-carrying bacteria in engineering applications and for understanding interspecies interactions in polymicrobial communities.

Alkylated cage silsesquioxane forming a long-range straight ordered hierarchical lamellar nanostructure.

An alkylated cage silsesquioxane (1), targeting for a new class of bottom-up-type fabricating materials, was successfully synthesized, and its self-assembled structure is described and discussed herein. Through this, it was found that the intermolecular interaction of long alkyl chains of 1 could be manipulated by thermal annealing to form a long-range straight ordered hierarchical lamellar structure with a periodicity of around 5 nm. Subsequent transmission electron microscopy (TEM) clearly identified polyhedral oligomeric silsesquioxane (POSS) molecules of 1 arranged in a highly ordered fashion, with a "head-to-head" type bilayered structure. The observation of a sublayer structure measuring approximately 0.4 nm in width was attributed to the highly regular packing of isobutyl groups in POSS molecules identified by TEM analysis. Moreover, the formation of a long-range straight structure with sharp interfacial boundaries, which is difficult to achieve with traditional diblock copolymers, is considered to be of significant importance to developing new practical applications of self-assembled nanostructures.