Mingwu Shen

PEGylated dendrimer-entrapped gold nanoparticles for in vivo blood pool and tumor imaging by computed tomography.

We report the synthesis and characterization of dendrimer-entrapped gold nanoparticles (Au DENPs) modified by polyethylene glycol (PEG) with enhanced biocompatibility for computed tomography (CT) imaging applications. In this study, amine-terminated poly(amidoamine) dendrimers of generation 5 (G5.NH(2)) modified by PEG monomethyl ether (G5.NH(2)-mPEG(20)) were used as templates to synthesize Au DENPs, followed by acetylation of the remaining dendrimer terminal amines to generate PEGylated Au DENPs. The partial PEGylation modification of dendrimer terminal amines allows high loading of Au within the dendrimer interior, and consequently by simply varying the Au salt/dendrimer molar ratio, the size of the PEGylated Au DENPs can be controlled at a range of 2-4 nm with a narrow size distribution. The formed PEGylated Au DENPs are water-dispersible, stable in a pH range of 5-8 and a temperature range of 0-50 °C, and non-cytotoxic at a concentration as high as 100 μm. X-ray absorption coefficient measurements show that the attenuation intensity of the PEGylated Au DENPs is much higher than that of Omnipaque with iodine concentration similar to Au. With the sufficiently long half-decay time demonstrated by pharmacokinetics studies, the PEGylated Au DENPs enabled not only X-ray CT blood pool imaging of mice and rats after intravenous injection of the particles, but also effective CT imaging of a xenograft tumor model in nude mice. These findings suggest that the designed PEGylated Au DENPs can be used as a promising contrast agent with enhanced biocompatibility for CT imaging of various biological systems, especially in cancer diagnosis.

Multifunctional dendrimer-entrapped gold nanoparticles for dual mode CT/MR imaging applications.

We report the synthesis, characterization, and utilization of gadolium-loaded dendrimer-entrapped gold nanoparticles (Gd-Au DENPs) for dual mode computed tomography (CT)/magnetic resonance (MR) imaging applications. In this study, amine-terminated generation five poly(amidoamine) dendrimers (G5.NH₂) modified with gadolinium (Gd) chelator and polyethylene glycol (PEG) monomethyl ether were used as templates to synthesize gold nanoparticles (AuNPs). Followed by sequential chelation of Gd(III) and acetylation of the remaining dendrimer terminal amine groups, multifunctional Gd-Au DENPs were formed. The formed Gd-Au DENPs were characterized via different techniques. We show that the formed Gd-Au DENPs are colloidally stable and non-cytotoxic at an Au concentration up to 50 μM. With the coexistence of two radiodense imaging elements of AuNPs and Gd(III) within one NP system, the formed Gd-Au DENPs display both r₁ relaxivity for MR imaging mode and X-ray attenuation property for CT imaging mode, which enables CT/MR dual mode imaging of the heart, liver, kidney, and bladder of rat or mouse within a time frame of 45 min. Furthermore, in vivo biodistribution studies reveal that the Gd-Au DENPs have an extended blood circulation time and can be cleared from the major organs within 24 h. The strategy to use facile dendrimer technology to design dual mode contrast agents may be extended to prepare multifunctional platforms for targeted multimode molecular imaging of various biological systems.

Hyaluronic acid-modified Fe3O4@Au core/shell nanostars for multimodal imaging and photothermal therapy of tumors

Development of multifunctional theranostic nanoplatforms for diagnosis and therapy of cancer still remains a great challenge. In this work, we report the use of hyaluronic acid-modified Fe3O4@Au core/shell nanostars (Fe3O4@Au-HA NSs) for tri-mode magnetic resonance (MR), computed tomography (CT), and thermal imaging and photothermal therapy of tumors. In our approach, hydrothermally synthesized Fe3O4@Ag nanoparticles (NPs) were used as seeds to form Fe3O4@Au NSs in the growth solution. Further sequential modification of polyethyleneimine (PEI) and HA affords the NSs with excellent colloidal stability, good biocompatibility, and targeting specificity to CD44 receptor-overexpressing cancer cells. With the Fe3O4 core NPs and the star-shaped Au shell, the formed Fe3O4@Au-HA NSs are able to be used as a nanoprobe for efficient MR and CT imaging of cancer cells in vitro and the xenografted tumor model in vivo. Likewise, the NIR absorption property enables the developed Fe3O4@Au-HA NSs to be used as a nanoprobe for thermal imaging of tumors in vivo and photothermal ablation of cancer cells in vitro and xenografted tumor model in vivo. This study demonstrates a unique multifunctional theranostic nanoplatform for multi-mode imaging and photothermal therapy of tumors, which may find applications in theranostics of different types of cancer.

Computed tomography imaging of cancer cells using acetylated dendrimer-entrapped gold nanoparticles

We report a new use of acetylated dendrimer-entrapped gold nanoparticles (Au DENPs) for in vitro and in vivo computed tomography (CT) imaging of cancer cells. In this study, Au DENPs prepared using amine-terminated generation 5 poly(amidoamine) dendrimers were subjected to an acetylation reaction to neutralize the positive surface potential. The acetylated Au DENPs were used for both in vitro and in vivo CT imaging of a human lung adencarcinoma cell line (SPC-A1 cells). Micro-CT images show that SPC-A1 cells can be detected under X-ray after incubation with the acetylated Au DENPs in vitro and the xenograft tumor model can be imaged after both intratumoral and intraperitoneal administration of the particles. Transmission electron microscopy data further confirm that the acetylated Au DENPs are able to be uptaken dominantly in the lysosomes of the cells. Combined morphological observation of cells after hematoxylin and eosin staining, MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay of cell viability, and flow cytometric analysis of cell cycle show that the acetylated Au DENPs do not appreciably affect the cell morphology, viability, and cell cycle, indicating their good biocompatibility at the given concentration range. Findings from this study suggest that the developed acetylated Au DENPs have a great potential to be used for CT imaging of cancer cells.

Encapsulation of 2-methoxyestradiol within multifunctional poly(amidoamine) dendrimers for targeted cancer therapy

We report here a general approach to using multifunctional poly(amidoamine) (PAMAM) dendrimer-based platform to encapsulate a potential anticancer drug for targeted cancer therapy. In this approach, amine-terminated generation 5 (G5) PAMAM dendrimers were sequentially modified with fluorescein isothiocyanate (FI) and folic acid (FA) via covalent conjugation, followed by an acetylation reaction to neutralize the remaining amines of the dendrimer surfaces. The synthesized multifunctional dendrimers (G5.NHAc-FI-FA) were then used to complex a potential anticancer drug, 2-methoxyestradiol (2-ME) for targeted delivery of the drugs to cancer cells overexpressing high-affinity folic acid receptors (FAR). We show that the formed G5.NHAc-FI-FA/2-ME complexes with each dendrimer encapsulating approximately 3.7 2-ME molecules are water soluble and stable. In vitro release studies show that 2-ME complexed with the multifunctional dendrimers can be released in a sustained manner. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in conjunction with cell morphology observation demonstrates that the G5.NHAc-FI-FA/2-ME complexes can specifically target and display specific therapeutic efficacy to cancer cells overexpressing high-affinity FAR. Findings from this study suggest that multifunctional dendrimers may be used as a general drug carrier to encapsulate various cancer drugs for targeted therapy of different types of cancer.

Electrospun poly(lactic-co-glycolic acid)/halloysite nanotube composite nanofibers for drug encapsulation and sustained release

We report a novel electrospun composite nanofiber-based drug delivery system. In this study, halloysite nanotubes (HNTs) were first used to encapsulate a model drug, tetracycline hydrochloride. Then, the drug-loaded HNTs with an optimized encapsulation efficiency were mixed with poly(lactic-co-glycolic acid) (PLGA) polymer for subsequent electrospinning to form drug-loaded composite nanofibrous mats. The structure, morphology, and mechanical properties of the formed electrospun composite nanofibrous mats were characterized using scanning electron microscopy (SEM), Fourier transform infrared spectroscopy, and tensile testing. In vitro drug release behavior was examined using UV-vis spectroscopy. The biocompatibility of HNT-containing PLGA fibers was evaluated through cell culture and MTT assay. We show that the incorporation of HNTs within the nanofibrous mats does not significantly change the morphology of the mats. In addition, our results indicate that this double-container drug delivery system (both PLGA polymer and HNTs are drug carriers) is beneficial to reduce the burst release of the drug and the introduction of HNTs can significantly improve the tensile strength of the polymer nanofibrous mats. Given the proved biocompatibility of the HNT-containing PLGA nanofibers via MTT assay of cell viability and SEM observation of cell morphology, the drug loaded electrospun composite nanofibrous mats developed in this study may find various applications in tissue engineering and pharmaceutical sciences.

Targeted CT/MR dual mode imaging of tumors using multifunctional dendrimer-entrapped gold nanoparticles

We report the synthesis and characterization of folic acid (FA)-modified multifunctional dendrimer-entrapped gold nanoparticles (Au DENPs) loaded with gadolinium (Gd) for targeted dual mode computed tomography (CT)/magnetic resonance (MR) imaging of tumors. In this work, amine-terminated generation 5 poly(amidoamine) dendrimers (G5.NH2) modified with Gd(III) chelator, polyethylene glycol (PEG) monomethyl ether, and PEGylated FA were used as templates to entrap gold nanoparticles (AuNPs). Further chelation of Gd(III) ions and acetylation of the remaining dendrimer terminal amines led to the formation of multifunctional FA-targeted Au DENPs loaded with Gd(III) (Gd-Au DENPs-FA). The formed Gd-Au DENPs-FA probes were characterized via different techniques. We show that the Gd-Au DENPs-FA probes with an Au NP core size of 4.0 nm are water dispersible, stable under different pH and temperature conditions, and cytocompatible in the given concentration range. With the co-existence of AuNPs and Gd(III) ions within the single multifunctional particles, Gd-Au DENPs-FA displayed high X-ray attenuation intensity and reasonable r1 relaxivity. These properties of the particles enabled them to be used as dual mode nanoprobes for targeted CT/MR imaging of cancer cells in vitro and xenograft tumor model in vivo via FA receptor-mediated active targeting pathway. The strategy to design multifunctional nanoprobes using the versatile dendrimer nanotechnology may be extended to design various dual mode or multimode imaging agents for accurate diagnosis of different types of cancer.

Polyethyleneimine-mediated synthesis of folic acid-targeted iron oxide nanoparticles for in vivo tumor MR imaging

We report a facile polyethyleneimine (PEI)-mediated approach to synthesizing folic acid (FA)-targeted magnetic iron oxide nanoparticles (Fe3O4 NPs) for in vivo magnetic resonance (MR) imaging of tumors. In this study, stable PEI-coated Fe3O4 NPs were prepared by a one-pot hydrothermal route. The aminated Fe3O4 NPs with PEI coating enabled covalent conjugation of fluorescein isothiocyanate (FI) and folate-conjugated polyethylene glycol (PEG) with one end of carboxyl groups (FA-PEG-COOH). Followed by final acetylation, FA-targeted PEGylated Fe3O4 NPs (Fe3O4-PEI-Ac-FI-PEG-FA NPs) were formed. The formed multifunctional Fe3O4 NPs were characterized via different techniques. We show that the PEI-mediated approach along with the PEGylation conjugation enables the generation of water-dispersible and stable multifunctional Fe3O4 NPs, and the particles are quite cytocompatible and hemocompatible in the given concentration range as confirmed by in vitro cytotoxicity assay, cell morphology observation, and hemolysis assay. In addition, flow cytometry and confocal microscopy data show that the multifunctional Fe3O4 NPs are able to target a model cancer cell line (KB cells) overexpressing FA receptors in vitro. Importantly, the FA-targeted Fe3O4 NPs are able to be used as an efficient nanoprobe for MR imaging of cancer cells in vitro and a xenografted tumor model in vivo via an active FA targeting pathway. With the facile PEI-mediated formation strategy and PEGylation conjugation chemistry, the Fe3O4 NPs may be multifunctionalized with other biological ligands for MR imaging of different biological systems.

Gene delivery using dendrimer-entrapped gold nanoparticles as nonviral vectors

Development of highly efficient nonviral gene delivery vectors still remains a great challenge. In this study, we report a new gene delivery vector based on dendrimer-entrapped gold nanoparticles (Au DENPs) with significantly higher gene transfection efficiency than that of dendrimers without AuNPs entrapped. Amine-terminated generation 5 poly(amidoamine) (PAMAM) dendrimers (G5.NH(2)) were utilized as templates to synthesize AuNPs with different Au atom/dendrimer molar ratios (25:1, 50:1, 75:1, and 100:1, respectively). The formed Au DENPs were used to complex two different pDNAs encoding luciferase (Luc) and enhanced green fluorescent protein (EGFP), respectively for gene transfection studies. The Au DENPs/pDNA polyplexes with different N/P ratios and compositions of Au DENPs were characterized by gel retardation assay, light scattering, zeta potential measurements, and atomic force microscopic imaging. We show that the Au DENPs can effectively compact the pDNA, allowing for highly efficient gene transfection into the selected cell lines as demonstrated by both Luc assay and fluorescence microscopic imaging of the EGFP expression. The transfection efficiency of Au DENPs with Au atom/dendrimer molar ratio of 25:1 was at least 100 times higher than that of G5.NH(2) dendrimers without AuNPs entrapped at the N/P ratio of 2.5:1. The higher gene transfection efficiency of Au DENPs is primarily due to the fact that the entrapment of AuNPs helps preserve the 3-dimensional spherical morphology of dendrimers, allowing for more efficient interaction between dendrimers and DNA. With the less cytotoxicity than that of G5.NH(2) dendrimers demonstrated by thiazoyl blue tetrazolium bromide assay and higher gene transfection efficiency, it is expected that Au DENPs may be used as a new gene delivery vector for highly efficient transfection of different genes for various biomedical applications.

Facile Hydrothermal Synthesis and Surface Functionalization of Polyethyleneimine-Coated Iron Oxide Nanoparticles for Biomedical Applications

We report the facile hydrothermal synthesis and surface functionalization of branched polyethyleneimine (PEI)-coated iron oxide nanoparticles (Fe3O4-PEI NPs) for biomedical applications. In this study, Fe3O4-PEI NPs were synthesized via a one-pot hydrothermal method in the presence of PEI. The formed Fe3O4-PEI NPs with primary amine groups on the surface were able to be further functionalized with polyethylene glycol (PEG), acetic anhydride, and succinic anhydride, respectively. The formed pristine and functionalized Fe3O4-PEI NPs were characterized via different techniques. We showed that the sizes of the Fe3O4-PEI NPs were able to be controlled by varying the mass ratio of Fe(II) salt and PEI. In addition, the formed Fe3O4-PEI NPs with different surface functionalities had good water dispersibility, colloidal stability, and relatively high R2 relaxivity (130-160 1/(mM·s)). Cell viability assay data revealed that the surface PEGylation and acylation of Fe3O4-PEI NPs rendered them with good biocompatibility in the given concentration range, while the pristine aminated Fe3O4-PEI NPs started to display slight toxicity at the concentration of 50 μg/mL. Importantly, macrophage cellular uptake results demonstrated that both PEGylation and acetylation of Fe3O4-PEI NPs were able to significantly reduce the nonspecific macrophage uptake, likely rendering the particles with prolonged circulation time. With the proven hemocompatibility and rich amine conjugation chemistry, the Fe3O4-PEI NPs with different surface functionalities may be applied for various biomedical applications, especially for magnetic resonance imaging and therapy.