Kasturi, SP, Sachaphibulkij, K and Roy, K. Covalent conjugation of polyethyleneimine on biodegradable microparticles for delivery of plasmid DNA vaccines. Biomaterials 26: 6375-6385

Department of Biomedical Engineering, The University of Texas at Austin, Austin, TX 78712, USA.
Biomaterials (Impact Factor: 8.56). 12/2005; 26(32):6375-85. DOI: 10.1016/j.biomaterials.2005.03.043
Source: PubMed


Microparticle-based delivery of nucleic acids has gained particular attention in recent years in view of improving the potency of DNA vaccination. Such improvement has been reported by encapsulation of pDNA within biodegradable microparticles or through surface adsorption on cationic microparticles. However, the intrinsic intracellular barriers for gene delivery to antigen presenting cells (APCs) have not been adequately addressed in the rational design of delivery systems for DNA vaccines. Here we report synthesis and characterization of biodegradable microparticles that (a) can passively target phagocytic APCs, (b) have intrinsic buffering ability that might allow for enhanced phagosomal escape, (c) are not cytotoxic and (d) have improved APC transfection efficiency. Branched polyethyleneimine (b-PEI) was covalently conjugated using carbodiimide chemistry to the surface of poly(lactide-coglycolide) (PLGA) microparticles to create cationic microparticles capable of simultaneously delivering both DNA vaccines as well as other immunomodulatory agents (cytokines or nucleic acids) within a single injectable delivery vehicle. Our results indicate that covalent conjugation of b-PEI allows efficient surface loading of nucleic acids, introduces intrinsic buffering properties to PLGA particles and enhances transfection of phagocytic cells without affecting the cytocompatibility of PLGA carriers.

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    • "Once inside the cell, these reagents can disrupt the endocytic pathway, releasing the contents of the endosomes to the cytosol.18,19 In fact, the covalent binding of PEI to the microparticle surface facilitates their incorporation into HeLa cells.14,20 However, the modification of microparticles with covalently bound PEI would render difficult or even impede their functionalization with other molecules, such as the drug to be delivered. "
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    ABSTRACT: Development of micro- and nanotechnology for the study of living cells, especially in the field of drug delivery, has gained interest in recent years. Although several studies have reported successful results in the internalization of micro- and nanoparticles in phagocytic cells, when nonphagocytic cells are used, the low internalization efficiency represents a limitation that needs to be overcome. It has been reported that covalent surface modification of micro- and nanoparticles increases their internalization rate. However, this surface modification represents an obstacle for their use as drug-delivery carriers. For this reason, the aim of the present study was to increase the capability for microparticle internalization of HeLa cells through the use of noncovalently bound transfection reagents: polyethyleneimine (PEI) Lipofectamine™ 2000 and FuGENE 6(®). Both confocal microscopy and flow cytometry techniques allowed us to precisely quantify the efficiency of microparticle internalization by HeLa cells, yielding similar results. In addition, intracellular location of microparticles was analyzed through transmission electron microscopy and confocal microscopy procedures. Our results showed that free PEI at a concentration of 0.05 mM significantly increased microparticle uptake by cells, with a low cytotoxic effect. As determined by transmission electron and confocal microscopy analyses, microparticles were engulfed by plasma-membrane projections during internalization, and 24 hours later they were trapped in a lysosomal compartment. These results show the potential use of noncovalently conjugated PEI in microparticle internalization assays.
    Full-text · Article · Nov 2012 · International Journal of Nanomedicine
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    • "In addition, the p-MPs can codeliver DNA and adjuvants generating an improved immune response [100, 101]. The current strategies have used polymers with cationic charge such polyethyleneimine to increase the loading and encapsulation efficiency of DNA inside particles [102]. Despite great advances in the development of DNA vaccines and their potential against several diseases, the biggest challenge is to establish the safety of using DNA vaccines in human medicine [103]. "
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    ABSTRACT: The development of the field of materials science, the ability to perform multidisciplinary scientific work, and the need for novel administration technologies that maximize therapeutic effects and minimize adverse reactions to readily available drugs have led to the development of delivery systems based on microencapsulation, which has taken one step closer to the target of personalized medicine. Drug delivery systems based on polymeric microparticles are generating a strong impact on preclinical and clinical drug development and have reached a broad development in different fields supporting a critical role in the near future of medical practice. This paper presents the foundations of polymeric microparticles based on their formulation, mechanisms of drug release and some of their innovative therapeutic strategies to board multiple diseases.
    Full-text · Article · May 2012 · BioMed Research International
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    • "Kasturi et al. found that pDNA encapsulated within PLGA leads to significant confinement of the complexes within endolysosomes, yet the application of chitosan to the surface of the NPs was projected to improve pDNA-NP release from the endolysosomes and offer a more protected path through the cytoplasm to the nucleus [18]. There are several reports on various aspects of chitosan- PLGA NPs for pDNA delivery, focusing on preparation methods, the final characteristics of synthesized NPs, and their behavior in vivo (i.e., cytotoxicity and transfection efficiency) [15–17, 19–21]. "
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    ABSTRACT: Poly(D,L-lactide-co-glycolide-) (PLGA-)chitosan nanoparticles are becoming an increasingly common choice for the delivery of nucleic acids to cells for various genetic manipulation techniques. These particles are biocompatible, with tunable size and surface properties, possessing an overall positive charge that promotes complex formation with negatively charged nucleic acids. This study examines properties of the PLGA-chitosan nanoparticle/plasmid DNA complex after formation. Specifically, the study aims to determine the optimal ratio of plasmid DNA:nanoparticles for nucleic acid delivery purposes and to elucidate the location of the pDNA within these complexes. Such characterization will be necessary for the adoption of these formulations in a clinical setting. The ability of PLGA-chitosan nanoparticles to form complexes with pDNA was evaluated by using the fluorescent intercalating due OliGreen to label free plasmid DNA. By monitoring the fluorescence at different plasmid: nanoparticle ratios, the ideal plasmid:nanoparticle ration for complete complexation of plasmid was determined to be 1:50. Surface-Enhanced Raman Spectroscopy and gel digest studies suggested that even at these optimal complexation ratios, a portion of the plasmid DNA was located on the outer complex surface. This knowledge will facilitate future investigations into the functionality of the system in vitro and in vivo.
    Full-text · Article · Jan 2011 · Journal of Nanomaterials
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