Jisang Yoo

Bioreducible branched poly(modified nona-arginine) cell-penetrating peptide as a novel gene delivery platform

Abstract Cell‐penetrating peptides (CPPs) have been widely used to deliver nucleic acid molecules. Generally, CPPs consisting of short amino acid sequences have a linear structure, resulting in a weak complexation and low transfection efficacy. To overcome these drawbacks, a novel type of CPP is required to enhance the delivery efficacy while maintaining its safe use at the same time. Herein, we report that a bioreducible branched poly‐CPP structure capable of responding to reducing conditions attained both outstanding delivery effectiveness and selective gene release in carcinoma cells. Branched structures provide unusually strong electrostatic attraction between DNA and siRNA molecules, thereby improving the transfection capability through a tightly condensed form. We designed a modified type of nona‐arginine (mR9) and synthesized a branched‐mR9 (B‐mR9) using disulfide bonds. A novel B‐mR9/pDNA polyplex exhibited redox‐cleavability and high transfection efficacy compared to conventional CPPs, with higher cell viability as well. B‐mR9/VEGF siRNA polyplex exhibited significant serum stability and high gene‐silencing effects in vitro. Furthermore, the B‐mR9 polyplex showed outstanding tumor accumulation and inhibition ability in vivo. The results suggest that the bioreducible branched poly CPP has great potential as a gene delivery platform. Graphical abstract Figure. No Caption available.

Establishment of a controlled insulin delivery system using a glucose-responsive double-layered nanogel

Glucose-responsive insulin delivery systems have been proposed as a promising alternative to conventional intramuscular administration methods, which causes low patient compliance due to the requirement of multiple administration. In addition, protein-based glucose-responsive systems using glucose oxidase and lectin have not achieved success in clinical trials because of their low biostability and potential cytotoxicity. In order to overcome these issues, the phenylboronic acid (PBA)-derivatives converted to hydrophilic moieties with an elevated glucose level play a key role in controlled insulin delivery systems due to their better biostability and high biocompatibility. In order to endow glucose-responsiveness to insulin delivery carriers using PBA derivatives, a glycol chitosan (GC)/sodium alginate (SA)-poly(L-glutmate-co-N-3-L-glutamylphenylboronic acid) (PGGA) graft polymer double-layered nanogel is synthesized by N-carboxyanhydride (NCA) polymerization and carbodiimide coupling reactions. The GC/SA-PGGA double-layered nanogel controllably releases insulin at diabetic glucose levels in vitro, and shows high biocompatibility, determined by cell viability and a hemolysis assay. Moreover, controlled insulin release at high glucose levels can be accomplished using the GC/SA-PGGA double-layered nanogel in mouse studies. Therefore, the GC/SA-PGGA double-layered nanogel characterized by glucose-sensitivity and superior biocompatibility may act as a potential platform for advanced insulin delivery systems.

Protease-activatable cell-penetrating peptide possessing ROS-triggered phase transition for enhanced cancer therapy

&NA; Reactive oxygen species (ROS)‐ or protease‐responsive materials have been utilized as carriers in cancer therapies because ROS and specific proteases are overproduced in cancer cells. Methionine‐based polypeptides containing a thioether group are promising candidates due to their ROS‐responsiveness which provides a phase transition. Herein, we developed protease‐activatable cell‐penetrating peptide containing a ROS‐responsive methionine, a cell permeable lysine chain, and a matrix metalloproteinase (MMP)‐cleavable linker. We designed a poly(l‐methionine‐block‐l‐lysine)‐PLGLAG‐PEG (MLMP) and doxorubicin (DOX) was loaded into the micelle core. The MLMP exhibited MMP‐sensitive cleavage and ROS‐induced DOX release. Moreover, we confirmed efficient DOX delivery into cancer cells and induction of the apoptotic capability in vitro. In a bio‐distribution study, IR‐780 dye encapsulated MLMP showed superior tumor targetability with long retention. Furthermore, MLMP (DOX) exhibited outstanding tumor inhibition capability with non‐toxicity compared to free DOX, indicating that dual stimuli‐MLMP has great potential as an anticancer drug delivery platform. Graphical abstract Figure. No caption available.

pH-controllable cell-penetrating polypeptide that exhibits cancer targeting

Helical peptides were naturally-occurring ordered conformations that mediated various biological functions essential for biotechnology. However, it was difficult for natural helical polypeptides to be applied in biomedical fields due to low bioavailability. To avoid these problems, synthetic alpha-helical polypeptides have recently been introduced by further modifying pendants in the side chain. In spite of an attractive biomimetic helical motif, these systems could not be tailored for targeted delivery mainly due to nonspecific binding events. To address these issues, we created a conformation-transformable polypeptide capable of eliciting a pH-activated cell-penetrating property solely at the cancer region. The developed novel polypeptide showed that the bare helical conformation had a function at physiological conditions while the pH-induced helical motif provided an active cell-penetrating characteristic at a tumor extracellular matrix pH. The unusual conformation-transformable system can elicit bioactive properties exclusively at mild acidic pH. STATEMENT OF SIGNIFICANCE We developed pH-controllable cell-penetrating polypeptides (PCCPs) undergoing pH-induced conformational transitions. Unlike natural cell-penetrating peptides, PCCPs was capable of penetrating the plasma membranes dominantly at tumor pH, driven by pH-controlled helicity. The conformation of PCCPs at neutral pH showed low helical propensity because of dominant electrostatic attractions within the side chains. However, the helicity of PCCPs was considerably augmented by the balance of electrostatic interactions, thereby inducing selective cellular penetration. Three polypeptides undergoing different conformational transitions were prepared to verify the selective cellular uptake influenced by their structures. The PCCP undergoing low-to-high helical conformation provided the tumor specificity and enhanced uptake efficiency. pH-induced conformation-transformable polypeptide might provide a novel platform for stimuli-triggered targeting systems.

Conformation-switchable helical polypeptide eliciting selective pro-apoptotic activity for cancer therapy

&NA; Artificial cationic helical peptides possess an enhanced cell‐penetrating property. However, their cell‐penetrability is not converted by cellular environmental changes resulting in nonspecific uptake. In this study, pH‐sensitive anion‐donating groups were added to a helical polypeptide to simultaneously achieve tumor targeting and pro‐apoptotic activity. The mitochondria‐destabilizing helical polypeptide undergoing pH‐dependent conformational transitions selectively targeted cancer cells consequently disrupting mitochondrial membranes and subsequently inducing apoptosis. This work presents a promising peptide therapeutic system for cancer therapy. Graphical abstract Mitochondria‐destabilizing helical polypeptide undergoing a pH‐activated conformational transition selectively perturbed the mitochondrial outer membranes thereby inducing pro‐apoptosis. Figure. No caption available.

Curcumin as a Novel Nanocarrier System for Doxorubicin Delivery to MDR Cancer Cells: In Vitro and In Vivo Evaluation

Curcumin (CRC) has been widely used as a therapeutic agent for various drug delivery applications. In this work, we focused on the applicability of CRC as a nanodrug delivery agent for doxorubicin hydrochloride (DOX) (commercially known as Adriamycin) coated with poly(ethylene glycol) (PEG) as an effective therapeutic strategy against multidrug-resistant cancer cells. The developed PEG-coated CRC/DOX nanoparticles (NPs) (PEG-CRC/DOX NPs) were well localized within the resistant cancer cells inducing apoptosis confirmed by flow cytometry and DNA fragmentation assays. The PEG-CRC/DOX NPs suppressed the major efflux proteins in DOX-resistant cancer cells. The in vivo biodistribution studies on HCT-8/DOX-resistant tumor xenograft showed improved bioavailability of the PEG-CRC/DOX NPs, and thereby suppressed tumor growth significantly compared to the other samples. This study clearly shows that curcumin nanoparticles could deliver DOX efficiently into the multidrug-resistant cancer cells to have potential therapeutic benefits.