Kang Moo Huh

In Vivo Biodistribution and Toxicology of Carboxylated Graphene Quantum Dots

Photoluminescent graphene quantum dots (GQDs) have fascinating optical and electronic properties with numerous promising applications in biomedical engineering. In this work, we first studied the in vivo biodistribution and the potential toxicity of carboxylated photoluminescent GQDs. KB, MDA-MB231, A549 cancer cells, and MDCK normal cell line were chosen as in vitro cell culture models to examine the possible adverse effects of the carboxylated photoluminescent GQDs. The carboxylated GQDs are desirable for increased aqueous solubility. All cancer cells efficiently took up the carboxylated GQDs. No acute toxicity or morphological changes were noted in either system at the tested exposure levels. A long-term in vivo study revealed that the GQDs mainly accumulated in liver, spleen, lung, kidney, and tumor sites after intravenous injection. To reveal any potential toxic effect of the GQDs on treated mice, serum biochemical analysis and histological evaluation were performed. The toxicity results from serum biochemistry and complete blood count study revealed that the GQDs do not cause appreciable toxicity to the treated animals. Finally, we observed no obvious organ damage or lesions for the GQDs treated mice after 21 days of administration at 5 mg/kg or 10 mg/kg dosages. With adequate studies of toxicity, both in vitro and in vivo, photoluminescent GQDs may be considered for biological application.

Biodegradable amphiphilic multiblock copolymers and their implications for biomedical applications

Alternating multiblock copolymers composed of short blocks of poly(ethylene oxide) (PEO) and poly(epsilon-caprolactone) (PCL) or poly(L-lactic acid) (PLLA) were synthesized by a coupling reaction. The block copolymers of relatively high molecular weights (M(n)20,000) formed a physically crosslinked thermoplastic network, while low molecular weight polymers were water-soluble. The block copolymers demonstrated solubility in a variety of solvents including acetone, tetrahydrofuran, methylene chloride, dioxane, water/acetone mixtures, and water/ethanol mixtures. The degree of swelling, optical transparency, and mechanical property of the films, prepared by a solvent casting method, were affected by the nature of the hydrophobic block used, polymer composition, temperature, and thermal history. The crystalline melting temperatures of PCL and PLLA in the block copolymers were significantly lowered due to the chemical structure of difunctional PCL and PLLA, and partial phase mixing with PEO segments. The properties of the block copolymers may be useful for biomedical applications as well as controlled drug release formulations. When PEO/PLLA multiblock copolymers were applied as a wound healing material loaded with basic fibroblast growth factor (bFGF), the feasibility study showed improved wound healing when compared to controls of no treatment and the same wound covering without bFGF, indicating that a certain degree of the bioactivity of bFGF is preserved.

Hydrotropic polymer micelles containing acrylic acid moieties for oral delivery of paclitaxel

Hydrotropic polymers (HPs) and their micelles have been recently developed as vehicles for delivery of poorly water-soluble drugs, such as paclitaxel (PTX), by oral administration. The release of PTX from HP micelles, however, was slow and it took more than a day for complete release of the loaded PTX. Since the gastrointestinal (GI) transit time is known to be only several hours, pH-sensitive HP micelles were prepared for fast release of the loaded PTX responding to pH changes along the GI tract. Acrylic acid (AA) was introduced, as a release modulator, into HPs by copolymerization with 4-(2-vinylbenzyloxy)-N,N-(diethylnicotinamide) (VBODENA). The AA content was varied from 0% to 50% (in the molar ratio to VBODENA). HPs spontaneously produced micelles in water, and their critical micelle concentrations (CMCs) ranged from 31 microg/mL to 86 microg/mL. Fluorescence probe study using pyrene showed that blank HP micelles possessed a good pH sensitivity, which was clearly observed at relatively high AA contents and pH>6. The pH sensitivity also affected the PTX loading property. Above pH 5, the PTX loading content and loading efficiency in HP micelles were significantly reduced. Although this may be primarily due to the AA moieties, other factors may include PTX degradation and polymer aggregation. The PTX release from HP micelles with more than 20% (mol) AA contents was completed within 12 h in a simulated intestinal fluid (SIF, pH=6.5). The HP micelles without any AA moiety showed very slow release profiles. In the simulated gastric fluid (SGF, pH=1.6), severe degradation of the released PTX was observed. The pH-dependent release of PTX from HP micelles can be used to increase the bioavailability of PTX upon oral delivery.

Targeted near-IR QDs-loaded micelles for cancer therapy and imaging

The use of water-soluble, functionalized quantum dots (QDs) that are highly stable against oxidation for biological and biomedical applications is currently one of the fastest growing fields of nanotechnology. Polymer-based nanoparticles are now widely used for drug delivery and targeted therapy. We modified the surface of near Infrared QDs by the solid dispersion method using PEG-PCDA and PCDA-Herceptin conjugates to demonstrate water-solubility and target-specific properties. Upon UV irradiation, QD cores located within nanoprobes were further stabilized by intramicellar cross-linking between neighboring PCDA-Herceptin moieties. These cross-linked nanoprobes showed higher stability and less toxicity. Near-IR QDs-loaded micelles were spherical with diameters of around 130-150 nm. The anti-tumor effect of near-IR QDs-loaded micelles against MDA-MB-231 tumors was remarkably better than that of control. Mice treated with the near-IR QDs-loaded micelles had a tumor volume of about 285 mm(3), indicating shrinkage in initial tumor volume and inhibition of tumor growth by 77.3% compared to that of control group (saline injection). In addition, near-IR QDs-loaded micelles were injected intravenously into tumor-bearing nude mice for simultaneous tumor therapy and imaging. We observed that the targeted near-IR QDs-loaded micelles distributed rapidly throughout the animal body including the tumor in real time. These multi-functional nanoprobes could therefore be used for both active and passive targeting, imaging and treatment of cancers in the early stage.

Cancer cell-specific photoactivity of pheophorbide a-glycol chitosan nanoparticles for photodynamic therapy in tumor-bearing mice

We designed a cancer-cell specific photosensitizer nano-carrier by synthesizing pheophorbide a (PheoA) conjugated glycol chitosan (GC) with reducible disulfide bonds (PheoA-ss-GC). The amphiphilic PheoA-ss-GC conjugates self-assembled in aqueous condition to form core-shell structured nanoparticles (PheoA-ss-CNPs) with good colloidal stability and switchable photoactivity. The photoactivity of PheoA-ss-CNPs in an aqueous environment was greatly suppressed by the self-quenching effect, which enabled the PheoA-ss-CNPs to remain photo-inactive and in a quenched state. However, after the cancer cell-specific uptake, the nanoparticular structure instantaneously dissociated by reductive cleavage of the disulfide linkers, followed by an efficient dequenching process. Compared to non-reducible PheoA-conjugated GC-NPs with stable amide linkages (PheoA-CNPs), PheoA-ss-CNPs rapidly restored their photoactivity in response to intracellular reductive conditions, thus presenting higher cytotoxicity with light treatment. In addition, the PheoA-ss-CNPs presented prolonged blood circulation in vivo compared to free PheoA, demonstrating enhanced tumor specific targeting behavior through the enhanced permeation and retention (EPR) effect. The enhanced tumor accumulation of PheoA-ss-CNPs enabled tumor therapeutic efficacy that was more efficient than free PheoA in tumor-bearing mice. Based on the enhanced intracellular release for cytosolic high dose and switchable photoactivity mechanism for reduced side effects, these results suggest that PheoA-ss-CNPs have good potential for photodynamic therapy (PDT) in cancer treatment.

GSH-mediated photoactivity of pheophorbide a-conjugated heparin/gold nanoparticle for photodynamic therapy.

In this study, we developed a new photosensitizer (PS)-conjugated hybrid nanoparticle comprised of gold nanoparticle (AuNP) as an efficient energy quencher, polysaccharide heparin and a second generation PS, pheophorbide a (PhA) for PDT. The hybrid nanoparticles (PhA-H/AuNPs) with an average size of 40nm were prepared by surface coating of AuNPs with PhA conjugated heparins via gold-thiol interaction. The glutathione (GSH)-mediated switchable photoactivity of the PhA-H/AuNPs was observed by fluorescence quenching and dequenching behaviors in the absence and presence of GSH. The photoactivity was significantly suppressed in aqueous media, but instantaneously restored at the GSH-rich intracellular environment to generate a strong fluorescence signal together with active production of singlet oxygen species with light treatment. In vitro cell tests revealed marked phototoxicity and high intracellular uptake of PhA-H/AuNPs in contrast with free PhA. The PhA-H/AuNPs also exhibited a prolonged circulation characteristic, enhanced tumor specificity, and improved photodynamic therapeutic efficacy compared with free PhA in tumor-bearing mice. As a result, the PhA-H/AuNPs may serve as an effective smart nanomedicine platform for PDT and have great potential for the clinical treatment of various tumors.

Synthesis and characterization of dextran grafted with poly(N‐isopropylacrylamide‐co‐N,N‐dimethyl‐acrylamide)

Graft copolymers consisting of dextran as a main chain and poly(N-isopropylacrylamide-co(N,N-dimethylacrylamide) (poly(NIPAAm-co-DMAAm)) as graft chains were synthesized. For the synthesis of the graft copolymers, a semitelechelic poly (NIPAAm-co-DMAAm) with an amino end-group was obtained by radical copolymerization with ethanethiol as a chain transfer agent, followed by a coupling reaction of its hydroxyl end-group with ethylenediamine. Graft copolymers with various length of the grafts were obtained from coupling reactions between carboxymethyl dextran and poly-(NIPAAm-co-DMAAm) in the presence of a water-soluble carbodiimide. The graft copolymers in phosphate buffer exhibit lower critical solution temperatures due to thermosensitivity of their grafts. There is no significant change in the hydration-dehydration behavior of the poly(NIPAAm-co-DMAAm) chain after the frafting reaction. The existence of such grafts in dextran may play an important role for modulated degradation in synchronization with temperature.

Biarmed Poly(ethylene glycol)-(pheophorbide a)2 Conjugate as a Bioactivatable Delivery Carrier for Photodynamic Therapy

In the study presented here, we developed a bioreducible biarmed methoxy poly(ethylene glycol)-(pheophorbide a)2 (mPEG-(ss-PhA)2) conjugate for cancer-cell-specific photodynamic therapy (PDT). PhA molecules were chemically conjugated with biarmed linkages at one end of the mPEG molecule via disulfide bonds. Under aqueous conditions, the amphiphilic mPEG-(ss-PhA)2 conjugate self-assembled to form core-shell-structured nanoparticles (NPs) with good colloidal stability. The mPEG-(ss-PhA)2 NPs exhibited intramolecular and intermolecular self-quenching effects that enabled the NPs to remain photoinactive in a physiological buffer. However, the dissociation of the NP structure was effectively induced by the cleavage of the disulfide bonds in response to intracellular reductive conditions, triggering the rapid release of PhA molecules in a photoactive form. In cell-culture systems, in addition to significant phototoxicity and intracellular uptake, we observed that the dequenching processes of PhA in the mPEG-(ss-PhA)2 NPs highly depended on the expression of intracellular thiols and that supplementation with glutathione monoethylester facilitated more rapid PhA release and enhanced the PhA phototoxicity. These findings suggest that the bioreducible activation mechanism of mPEG-(ss-PhA)2 NPs in cancer cells can maximize the cytosolic dose of active photosensitizers to achieve high cytotoxicity, thereby enhancing the treatment efficacy of photodynamic cancer treatment.

Preparation and characterization of glycol chitin as a new thermogelling polymer for biomedical applications.

In this study, a new thermo-sensitive polymer, glycol chitin, was synthesized by controlled N-acetylation of glycol chitosan and evaluated as a thermogelling system. The physico-chemical properties of glycol chitins with different degrees of acetylation (DA) were investigated in terms of degradation, cytotoxicity, rheological properties, and in vitro and in vivo gel formation. Aqueous solutions of glycol chitins were flowable freely at room temperature but quickly became a durable gel at body temperature. Thermo-reversible sol-gel transition properties were observed with fast gelation kinetics. Glycol chitins with higher DA showed faster degradation in the presence of lysozyme. They exhibited no significant biological toxicity against human cell lines. An anti-cancer drug, doxorubicin, could be incorporated into the hydrogel by a simple mixing process and released in a sustained pattern over 13 days. Our findings suggest that glycol chitins could be useful as a new thermogelling biomaterial for drug delivery and injectable tissue engineering.

Encapsulation of CdSe/ZnS Quantum Dots in Poly(ethylene glycol)-Poly(D,L-Lactide) Micelle for Biomedical Imaging and Detection

Luminescent CdSe/ZnS QDs, with emission in the red region of the spectrum, were synthesized and encapsulated in poly(ethylene glycol)-poly(D,L-lactide) diblock copolymer micelles, to prepare water-soluble, biocompatible QD micelles. PEG-PLA diblock copolymers were synthesized by ring opening polymerization of D,Llactide, in the presence of methoxy PEG as a macroinitiator. QDs were encapsulated with PEG-PLA polymers using a solid dispersion method in chloroform. The resultant polymer micelles, with encapsulated QDs, were characterized using various analytical techniques, such as UV-Vis measurement, light scattering, fluorescence spectroscopy, transmission electron microscopy (TEM) and atomic forced microscopy (AFM). The polymer micelles, with encapsulated QDs, were spherical and showed diameters in the range of 20–150 nm. The encapsulated QDs were highly luminescent, and have high potential for applications in biomedical imaging and detection.