Induced packaging of mRNA into polyplex micelles by regulated hybridization with a small number of cholesteryl RNA oligonucleotides directed enhanced in vivo transfection.

There has been a progressive interest in the molecular design of polymers and lipids as synthetic carriers for targeting therapeutic mRNA in vivo with the ability to circumvent nuclease attack for treating intractable diseases. Herein, we developed a simple approach to attain one order of magnitude higher nuclease tolerability of mRNA through the formation of polyplex micelles (PMs) by combining ω-cholesteryl (ω-Chol)-poly (ethylene-glycol) (PEG)-polycation block copolymers with mRNA pre-hybridized with cholesterol (Chol)-tethered RNA oligonucleotides (Chol (+)-OligoRNA). Even one or a few short Chol (+)-OligoRNA anchors harboring along the 46-fold longer mRNA strand was sufficient to induce tight mRNA packaging in the PM core, as evidenced by Förster resonance energy transfer (FRET) measurement as well as by a longitudinal relaxation time (T) measurement using NMR.

Design concepts of polyplex micelles for in vivo therapeutic delivery of plasmid DNA and messenger RNA.

Nonviral delivery of plasmid (p)DNA or messenger (m)RNA is a safe and promising therapeutic option to continuously supply therapeutic proteins into diseased tissues. In most cases of in vivo pDNA and mRNA delivery, these nucleic acids are loaded into carriers based on cationic polymers and/or lipids to prevent nuclease-mediated degradation before reaching target cells. The carriers should also evade host clearance mechanisms, including uptake by scavenger cells and filtration in the spleen.

Glucose-linked sub-50-nm unimer polyion complex-assembled gold nanoparticles for targeted siRNA delivery to glucose transporter 1-overexpressing breast cancer stem-like cells.

Cancer stem-like cells (CSCs) treatment is a plausible strategy for enhanced cancer therapy. Here we report a glucose-installed sub-50-nm nanocarrier for the targeted delivery of small interfering RNA (siRNA) to CSCs through selective recognition of the glucose ligand to the glucose transporter 1 (GLUT1) overexpressed on the CSC surface. The siRNA nanocarrier was constructed via a two-step assembling process.

Poly( N-isopropylacrylamide)-Based Polymer-Inducing Isothermal Hydrophilic-to-Hydrophobic Phase Transition via Detachment of Hydrophilic Acid-Labile Moiety.

The polymerization of N-isopropylacrylamide (NIPAAm) with ionizable monomers results in pH-responsive lower critical solution temperature (LCST) polymer which works in an ionization-dependent manner. However, gradual ionization of the comonomer occurs at a broad pH range due to the electrostatic field generated by the polymers, limiting the extent of LCST shift in response to pH change. Furthermore, excess introduction of comonomer may dull phase transition behavior.

Precise tuning of disulphide crosslinking in mRNA polyplex micelles for optimising extracellular and intracellular nuclease tolerability.

The major issues in messenger (m)RNA delivery are rapid mRNA degradation in the extracellular and intracellular spaces, which decreases the efficiency and duration for protein expression from mRNA. Stabilization of mRNA carriers using environment-responsive crosslinkings has promises to overcome these issues. Herein, we fine-tuned the structure of disulphide crosslinkings, which are selectively cleaved in the intracellular reductive environment, using the mRNA-loaded polyplex micelles (PMs) prepared from poly(ethylene glycol)-poly(L-lysine) (PEG-PLys) block copolymers, particularly by focussing on cationic charge density after the crosslinking.

Epirubicin-loaded polymeric micelles effectively treat axillary lymph nodes metastasis of breast cancer through selective accumulation and pH-triggered drug release.

Suppression of axillary lymph node metastasis (ALNM) is an important goal in the treatment of breast cancer. While several therapies directed to ALNM have been evaluated, effective and safe treatments for ALNM in triple negative breast cancer (TNBC) have not been established yet, especially against initial/small metastases. Here, we demonstrated the therapeutic effect of an anthracycline drug, epirubicin (EPI)-loaded polymeric micelles equipped with pH-triggered drug release property (EPI/m) against ALNM of TNBC.

Robust Polyion Complex Vesicles (PICsomes) under Physiological Conditions Reinforced by Multiple Hydrogen Bond Formation Derived by Guanidinium Groups.

Polyion complex vesicles (PICsomes) formed from a self-assembly of an oppositely charged pair of block- and homo-polyelectrolytes have shown exceptional features for functional loading of bioactive agents. Nevertheless, the stability of PICsomes is often jeopardized in a physiological environment, and only PICsomes having chemically cross-linked membranes have endured in harsh in vivo conditions, such as in the bloodstream. Herein, we developed versatile PICsomes aimed to last in in vivo settings by stabilizing their membrane through a combination of ionic and hydrogen bonding, which is widely found in natural proteins as a salt bridge, by controlled introduction of guanidinium groups in the polycation fraction toward concurrent polyion complexation and hydrogen bonding.

Treatment of Bone Defects by Transplantation of Genetically Modified Mesenchymal Stem Cell Spheroids.

Cell transplantation is promising for regenerative medicine. A combination of a three-dimensional spheroid culture system with gene transfection was developed to enhance the therapeutic effects of mesenchymal stem cell (MSC) transplantation. The spheroid cell culture system is based on micropatterned substrates composed of a regular array of 100-μm-diameter cell-adhesion areas coated with a temperature-responsive polymer, poly (N-isopropylacrylamide-co-methacrylic acid), which allows for spheroid detachment by simply cooling the plates.

Prolonged engraftment of transplanted hepatocytes in the liver by transient pro-survival factor supplementation using ex vivo mRNA transfection.

Cell transplantation therapy needs engraftment efficiency improvement of transplanted cells to the host tissues. Ex vivo transfection of a pro-survival gene to transplanted cells is a possible solution; however prolonged expression and/or genomic integration of the gene can be cancer promoting. To supply pro-survival protein only when it is needed, we used mRNA transfection, which exhibits transient protein expression profiles without the risk of genomic integration.

Block Copolymer Micelles in Nanomedicine Applications.

Polymeric micelles are demonstrating high potential as nanomedicines capable of controlling the distribution and function of loaded bioactive agents in the body, effectively overcoming biological barriers, and various formulations are engaged in intensive preclinical and clinical testing. This Review focuses on polymeric micelles assembled through multimolecular interactions between block copolymers and the loaded drugs, proteins, or nucleic acids as translationable nanomedicines. The aspects involved in the design of successful micellar carriers are described in detail on the basis of the type of polymer/payload interaction, as well as the interplay of micelles with the biological interface, emphasizing on the chemistry and engineering of the block copolymers.

Effect of multiple cyclic RGD peptides on tumor accumulation and intratumoral distribution of IRDye 700DX-conjugated polymers.

Strategic delivery of IRDye 700DX (photosensitizer) is a key for improving its effect in photodynamic therapy. In this study, we have synthesized IRDye 700DX-conjugated polymers containing multiple cyclic RGD peptides to deliver IRDye 700DX selectively to tumor cells and tumor-associated blood vessels overexpressing αβ integrin. Our polymer has a backbone of hydrophilic poly(ethylene glycol)-poly(L-glutamic acid) block copolymer, and cyclic RGD peptides are conjugated to side chains of the poly(L-glutamic acid) while IRDye 700DX is conjugated to the terminal of poly(ethylene glycol).

Tuned Density of Anti-Tissue Factor Antibody Fragment onto siRNA-Loaded Polyion Complex Micelles for Optimizing Targetability into Pancreatic Cancer Cells.

Antibody fragment (Fab')-installed polyion complex (PIC) micelles were constructed to improve targetability of small interfering RNA (siRNA) delivery to pancreatic cancer cells. To this end, we synthesized a block copolymer of azide-functionalized poly(ethylene glycol) and poly(l-lysine) and prepared PIC micelles with siRNA. Then, a dibenzylcyclooctyne (DBCO)-modified antihuman tissue factor (TF) Fab' was conjugated to azido groups on the micellar surface.

Combined CatWalk Index: an improved method to measure mouse motor function using the automated gait analysis system.

Objective: Measuring motor function in mice is important for studying models of spinal cord injury (SCI) or other diseases. Several methods exist based on visual observation of mice moving in an open field. Though these methods require very little equipment, observers must be trained, and the possibility of human error or subjectivity cannot be eliminated.

An Ethylenediamine-based Switch to Render the Polyzwitterion Cationic at Tumorous pH for Effective Tumor Accumulation of Coated Nanomaterials.

Polyzwitterions are employed as coating polymers for biomaterials to induce an antifouling property on the surface. Fine-tuning the betaine structure switches the antifouling property to be interactive with anionic tissue constituents in response to a tumorous pH gradient. The ethylenediamine moiety in the carboxybetaine enabled stepwise protonation and initiated the di-protonation process around tumorous pH (6.

Enhanced Intracellular Delivery of siRNA by Controlling ATP-Responsivity of Phenylboronic Acid-Functionalized Polyion Complex Micelles.

Intracellular delivery of small interfering RNA (siRNA) is a long-standing challenge in oligonucleotide therapeutics. Herein, adenosine triphosphate (ATP)-responsive polyion complex micelles assembled from poly(ethylene glycol)-block-poly(l-lysine) (PEG-PLys) bearing 4-carboxy-3-fluorophenylboronic acid (FPBA) moiety in the PLys side chains (FPBA micelle) for the delivery of cholesterol-modified siRNA (Chol-siRNA) are described. The pK of FPBA moiety is 7.

Synthetic "smart gel" provides glucose-responsive insulin delivery in diabetic mice.

Although previous studies have attempted to create "electronics-free" insulin delivery systems using glucose oxidase and sugar-binding lectins as a glucose-sensing mechanism, no successful clinical translation has hitherto been made. These protein-based materials are intolerant of long-term use and storage because of their denaturing and/or cytotoxic properties. We provide a solution by designing a protein-free and totally synthetic material-based approach.

Polyplex Micelles with Phenylboronate/Gluconamide Cross-Linking in the Core Exerting Promoted Gene Transfection through Spatiotemporal Responsivity to Intracellular pH and ATP Concentration.

Polyplexes as gene delivery carriers require integrated functionalities to modulate intracellular trafficking for efficient gene transfection. Herein, we developed plasmid DNA (pDNA)-loaded polyplex micelles (PMs) from poly(ethylene glycol)-based block catiomers derivatized with 4-carboxy-3-fluorophenylboronic acid (FPBA) and d-gluconamide to form pH- and ATP-responsive cross-linking in the core. These PMs exhibited robustness in the extracellular milieu and smooth endosomal escape after cellular uptake, and they facilitated pDNA decondensation triggered by increased ATP concentration inside of the cell.

Glycaemic control boosts glucosylated nanocarrier crossing the BBB into the brain.

Recently, nanocarriers that transport bioactive substances to a target site in the body have attracted considerable attention and undergone rapid progression in terms of the state of the art. However, few nanocarriers can enter the brain via a systemic route through the blood-brain barrier (BBB) to efficiently reach neurons. Here we prepare a self-assembled supramolecular nanocarrier with a surface featuring properly configured glucose.

Designing immunostimulatory double stranded messenger RNA with maintained translational activity through hybridization with poly A sequences for effective vaccination.

Messenger (m)RNA vaccines require a safe and potent immunostimulatory adjuvant. In this study, we introduced immunostimulatory properties directly into mRNA molecules by hybridizing them with complementary RNA to create highly immunogenic double stranded (ds)RNAs. These dsRNA formulations, comprised entirely of RNA, are expected to be safe and highly efficient due to antigen expression and immunostimulation occurring simultaneously in the same antigen presenting cells.

Therapeutic Vesicular Nanoreactors with Tumor-Specific Activation and Self-Destruction for Synergistic Tumor Ablation.

Polymeric nanoreactors (NRs) have distinct advantages to improve chemical reaction efficiency, but the in vivo applications are limited by lack of tissue-specificity. Herein, novel glucose oxidase (GOD)-loaded therapeutic vesicular NRs (theraNR) are constructed based on a diblock copolymer containing poly(ethylene glycol) (PEG) and copolymerized phenylboronic ester or piperidine-functionalized methacrylate (P(PBEM-co-PEM)). Upon systemic injection, theraNR are inactive in normal tissues.

Functional polymer-based siRNA delivery carrier that recognizes site-specific biosignals.

Responsive molecular designs to specific biosignals in microenvironments endow site-specific functionalities with associated polymers. Thus, the construction of small interfering RNA (siRNA) carriers with functional polymers enables smart programs that are triggered by sequential biosignals in a pathway to the targeted cytosol for effective gene silencing. In this review, we explain rational strategies for the design of functional polymers with responsiveness to biosignals and describe the examples of smart carriers for siRNA delivery.

Effective treatment of drug resistant recurrent breast tumors harboring cancer stem-like cells by staurosporine/epirubicin co-loaded polymeric micelles.

Breast cancer recurrence and resistance are associated with cancer stem-like cell (CSC) sub-populations. As conventional therapies fail to treat CSCs, institution of novel therapeutic strategies capable of eradicating both cancer cells and CSCs is central for achieving effective treatments with long-term survival. Here, we studied the ability of polymeric micelles cooperatively loading the cytotoxic drug epirubicin (Epi) and the CSC inhibitor staurosporine (STS) to treat breast tumors, particularly when tumors relapsed after chemotherapy.

Engineering Tumour Cell-Binding Synthetic Polymers with Sensing Dense Transporters Associated with Aberrant Glutamine Metabolism.

Increased glutamine uptake toward the elevated glutaminolysis is one of the hallmarks of tumour cells. This aberrant glutamine metabolism has recently attracted considerable attention as a diagnostic and therapeutic target. Herein, we developed glutamine-functionalized polymer to achieve a selective high affinity to tumour cells overexpressing glutaminolysis-related transporter ASCT2.

Nanomaterial-Enabled Cancer Therapy.

While cancer remains the major cause of death worldwide, nanomaterial (NM)-based diagnosis and treatment modalities are showing remarkable potential to better tackle clinical oncology by effectively targeting therapeutic agents to tumors. NMs can selectively accumulate in solid tumors, and they can improve the bioavailability and reduce the toxicity of encapsulated cytotoxic agents. Additional noteworthy functions of NMs in cancer treatment include the delivery of contrast agents to image tumor sites, delivery of genetic materials for gene therapy, and co-delivery of multiple agents to achieve combination therapy or simultaneous diagnostic and therapeutic outcomes.