Hyun Su Min

pH-Controlled Gas-Generating Mineralized Nanoparticles: A Theranostic Agent for Ultrasound Imaging and Therapy of Cancers

We report a theranostic nanoparticle that can express ultrasound (US) imaging and simultaneous therapeutic functions for cancer treatment. We developed doxorubicin-loaded calcium carbonate (CaCO3) hybrid nanoparticles (DOX-CaCO3-MNPs) through a block copolymer templated in situ mineralization approach. The nanoparticles exhibited strong echogenic signals at tumoral acid pH by producing carbon dioxide (CO2) bubbles and showed excellent echo persistence. In vivo results demonstrated that the DOX-CaCO3-MNPs generated CO2 bubbles at tumor tissues sufficient for echogenic reflectivity under a US field. In contrast, the DOX-CaCO3-MNPs located in the liver or tumor-free subcutaneous area did not generate the CO2 bubbles necessary for US contrast. The DOX-CaCO3-MNPs could also trigger the DOX release simultaneously with CO2 bubble generation at the acidic tumoral environment. The DOX-CaCO3-MNPs displayed effective antitumor therapeutic activity in tumor-bearing mice. The concept described in this work may serve as a useful guide for development of various theranostic nanoparticles for US imaging and therapy of various cancers.

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.

Nanobubbles from Gas-Generating Polymeric Nanoparticles: Ultrasound Imaging of Living Subjects†

Ultrasound (US) is a noninvasive biomedical imaging modality that is widely available, inexpensive, safe, and provides real-time imaging and diagnosis. US contrast enhancement improves the image quality because of acoustic impedance mismatch, which can occur when a liquid or gas is encapsulated within shell materials or when liquid or solid particles with a high density difference are brought together. Microbubbles were primarily used as US contrast agents for blood flow imaging, however, development of metabolic or molecular US imaging that employs microbubbles has been limited because sub-microbubbles or nanoparticles that are appropriate for tissue penetration have poor echogenic sensitivity, while nanoparticles can not be visualized at the resolution of US imaging instruments (typically 50 to 100 mm). Recently, nanoparticles have been employed in numerous biomedical applications. Nanoparticles bypass the reticuloendothelial system, and allow prolonged circulation and passive localization within tumor vasculature by tissue extravasation and enhanced permeation and retention (EPR). Moreover, nanoparticles can be modified by addition of ligands that increase nanoparticle affinity toward specific target sites, such as tumors. In addition, the insertion of protease cleavage sites can provide activatable imaging probes, thus allowing simultaneous molecular imaging and therapeutics. The need to use nanosized carriers for in vivo imaging and therapy has led to the development of laser-induced photoacoustic US and to the exploration of materials-based approaches that employ perfluorocarbonencapsulated lipid-based nanoparticles and polymeric micelles. Herein, we describe the generation of nanobubbles, which can be imaged by US and are derived from gas-generating polymeric nanoparticles (GGPNPs). The proposed mechanism involves localization of echogenic GGPNPs in a tumor and coalescence of generated nanobubbles, followed by fusion of nanobubbles into microbubbles. Significantly, we produced GGPNP nanobubbles without encapsulation of a gas precursor (perfluorocarbon). Instead, in our system, the GGPNP carbonate side chain is degraded to form carbon dioxide. The mechanism appears to involve diffusion of water into the GGPNPs, cleavage of the carbonate side chains by hydrolysis, and formation of carbon dioxide nanobubbles on the GGPNP surface, followed by expansion or coalescence of nanobubbles into microbubbles. The resulting microbubbles exhibit resonance under a US field (Figure 1a). Polyesters with carbonate side chains (poly(BL-PO)) were synthesized by ring-opening copolymerization of gbutyrolactone and propylene oxide using samarium diiodide as initiator (Figure S1b in the Supporting Information). The carbonate side chains were created by conjugation with hydroxy groups of hydroxy g-butyrolactone and cholesteryl chloroformate or ethyl chloroformate (Figure S1–S3 in the Supporting Information). The copolymerization ratios of cholesteryl carbonate g-butyrolactone and ethyl carbonate g-butyrolactone were varied with respect to that of propylene oxide. The resulting chemical structures are shown in Figure 1b. The initial feed ratios of three monomers were 1:2:1 for poly(BL-PO). The actual ratio of three monomer units in the poly(BL-PO) was confirmed as 1:1.74:0.81 by H NMR spectroscopy (see Figure S4 in the Supporting Information). This result shows that the compositions of the copolymers were well-matched to the initial feed ratio. The presence of the cholesteryl group means that the relative proportions of cholesteryl carbonate and ethyl carbonate determine the physical stability of the polymer. The hydrolyzed particles produce biocompatible water, carbon dioxide, cholesterol, and ethanol. To prove the biocompatibility of our particles, we examined their effect on cell viability. At a concentration of up to 1 mgmL , our particles were not cytotoxic (see Figure S8 in the Supporting Information). Nanoparticles generated from poly(BL-PO) (1:2:1) were termed GGPNPs. Dynamic light scattering (DLS) indicated that the average GGPNP diameter was 581 nm (Figure S6 in the Supporting Information), and transmission electron microscopy (TEM) indicated that the GGPNP morphology was spherical and the diameter was less than 500 nm (Figure 1c). The size appeared larger by DLS probably [*] Dr. E. Kang, H. S. Min, Dr. H. J. Ahn, Dr. I. C. Yoon, Dr. K. Choi, Dr. K. Kim, Dr. I. C. Kwon Biomedical Research Center Korea Institute of Science and Technology 39-1 Hawolgok-dong, Seongbuk-gu, Seoul 136 791 (Korea) Fax: (+82)2-958-5909 E-mail: ikwon@kist.re.kr

Tumor-targeting glycol chitosan nanoparticles as a platform delivery carrier in cancer diagnosis and therapy.

A natural based polymer, chitosan has received widespread attention in drug delivery systems due to its valuable physicochemical and biological characteristics. In particular, hydrophobic moiety-conjugated glycol chitosan can form amphiphilic self-assembled glycol chitosan nanoparticles (GCNPs) and simultaneously encapsulate hydrophobic drug molecules inside their hydrophobic core. This GCNP-based drug delivery systems exhibit excellent tumor-homing efficacy, attributed to the long blood circulation and the enhanced permeability and retention effect; this tumor-targeting drug delivery results in improved therapeutic efficiency. In this review, we describe the requisite properties of GCNPs for cancer therapy as well as imaging for diagnosis, such as their basic characteristics, in vitro delivery efficiency and in vivo tumor-targeting ability.

Echogenic Glycol Chitosan Nanoparticles for Ultrasound-Triggered Cancer Theranostics.

Theranostic nanoparticles hold great promise for simultaneous diagnosis of diseases, targeted drug delivery with minimal toxicity, and monitoring of therapeutic efficacy. However, one of the current challenges in developing theranostic nanoparticles is enhancing the tumor-specific targeting of both imaging probes and anticancer agents. Herein, we report the development of tumor-homing echogenic glycol chitosan-based nanoparticles (Echo-CNPs) that concurrently execute cancer-targeted ultrasound (US) imaging and US-triggered drug delivery. To construct this novel Echo-CNPs, an anticancer drug and bioinert perfluoropentane (PFP), a US gas precursor, were simultaneously encapsulated into glycol chitosan nanoparticles using the oil in water (O/W) emulsion method. The resulting Echo-CNPs had a nano-sized particle structure, composing of hydrophobic anticancer drug/PFP inner cores and a hydrophilic glycol chitosan polymer outer shell. The Echo-CNPs had a favorable hydrodynamic size of 432 nm, which is entirely different from the micro-sized core-empty conventional microbubbles (1-10 μm). Furthermore, Echo-CNPs showed the prolonged echogenicity via the sustained microbubble formation process of liquid-phase PFP at the body temperature and they also presented a US-triggered drug release profile through the external US irradiation. Interestingly, Echo-CNPs exhibited significantly increased tumor-homing ability with lower non-specific uptake by other tissues in tumor-bearing mice through the nanoparticles enhanced permeation and retention (EPR) effect. Conclusively, theranostic Echo-CNPs are highly useful for simultaneous cancer-targeting US imaging and US-triggered delivery in cancer theranostics.