Evaluation of transferrin-polyethylenimine conjugate for targeted gene delivery.
ABSTRACT With the aim to improve the specificity and to reduce the cytotoxicity of polyethylenimine (PEI), we have synthesized the conjugates of the branched PEI (25 kDa) with transferrin. The transferrin-PEI (TP) conjugates with five compositions were synthesized using periodate oxidation method and confirmed by FT-IR spectroscopy and gel permeation chromatography. The free amine contents of TP conjugates, which were able to condense and deliver DNA, increased as the amount of PEI increased. TP/DNA polyplexes were characterized by measuring gel electrophoresis, ethidium bromide fluorescence quenching, particle size and zeta potential of complexes. Complete complexation of the polyplexes was observed above the N/P ratio of 5 in TP/ DNA, and above 3 in PEI/DNA, respectively. The zeta potential of the complexes decreased as the amount of transferrin in TP conjugates increased. Transfection efficiency of TP conjugates was evaluated in HeLa cell and Jurkat cell systems. Among the five compositions of TP conjugates, TP-2 system mediated a higher beta-galactosidase gene expression than PEI system in Jurkat cell which was known to express elevated numbers of transferrin receptors. From the results of the cell viability based on MTT assay, TP conjugates showed lower cytotoxicity compared with the PEI system. We expect that the TP conjugate can be used efficiently as a nonviral gene delivery vector.
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ABSTRACT: Cisplatin is a major broad-spectrum chemotherapeutic agent, however, its dose-dependent side effects limit the administration of large doses. Presently, developing a drug targeted delivery system is suggested as one of the most promising approaches to minimize the side effects of cisplatin. Here, we found that each human serum transferrin (HTf) has the potential to bind with over 22 cisplatins, and the complex of apo-HTf-cisplatin can specifically deliver cisplatin to HepG2 cells (human hepatocellular liver carcinoma cell line) in vitro, and facilitate HepG2 cells to apoptosis. Moreover, proteomics methods revealed that the abundances of 23 proteins in HepG2 cells were remarkably altered in response to cisplatin/apo-HTf-cisplatin exposure, and Realtime-PCR revealed that a number of important genes related to chemotherapeutic cytotoxicity and chemotherapeutic resistance are differentially transcribed between the HepG2 cells of cisplatin exposed and HTf-cisplatin exposed. The pathway analysis of the differentially expressed proteins and gene transcriptions indicated that those regulated proteins and gene transcriptions are involved in apoptosis regulation, transcription, cell cycle control, protein biosynthesis, energy metabolism, signal transduction, protein binding and other functions. It indicated that the cisplatin toxicity in HepG2 cell is diverse, the transport process has an effect on the cisplatin cytotoxicity, and the mechanism of the apoptosis of HepG2 cells induced by apo-HTf-cisplatin is different from that of cisplatin.Journal of proteomics 09/2012; · 5.07 Impact Factor
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ABSTRACT: In this study lactoferrin (Lf) was investigated as a targeting ligand for receptor-mediated gene delivery to human bronchial epithelial cells. A high number of lactoferrin receptors (LfRs) were detected on bronchial epithelial (BEAS-2B), but not on alveolar epithelial (A549) cells by fluorescence microscopy and FACS measurements, suggesting potential targeting selectivity for bronchial epithelial cells. Molecular conjugates with ratios of Lf to branched polyethylenimine 25 kDa (PEI) ranging from 4:1 to 1:40 (mol/mol) were synthesized and analyzed for complexation of plasmid DNA (pDNA), transfection efficiency, and cytotoxicity. Whereas particle size increased with the degree of Lf coupling from 45 to 225 nm, surface charge was not significantly influenced. Transfection studies on BEAS-2B cells revealed that Lf-PEI 1:20 exhibited the highest luciferase gene expression which was 5-fold higher at an N/P ratio (molar ratio of PEI nitrogen to pDNA phosphate) of 4 than PEI and could be inhibited by an excess of free Lf. With A549 cells, no significant enhancement in transfection efficiency between Lf-PEI/pDNA and PEI/pDNA complexes could be observed. Increasing the degree of Lf coupling to PEI resulted in reduced transfection efficiency in both alveolar and bronchial epithelial cells. Cell viability assays resulted in significantly lower cellular toxicity of Lf-PEI/pDNA compared with PEI/pDNA complexes. We suggest that Lf represents a potent targeting ligand for receptor-mediated gene delivery to bronchial epithelial cells and might be a promising candidate for lung gene transfer in vivo.Biomaterials 09/2007; 28(23):3448-55. · 8.31 Impact Factor
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ABSTRACT: For a new anticancer drug carrier, we synthesized 4 compositions of amphiphilic stearic acidconjugated pullulan (SAP) and characterized them with FT-IR spectroscopy. Crystalline changes were verified by x-ray diffraction patterns before and after synthesis of the SAP conjugate. SAP nanoparticles were prepared by a diafiltration method, and the fluorescence spectroscopy using pyrene showed particle self-assembly in water. SAP nanoparticles were spherical in TEM photos, and particle size ranged between 200 approximately 500 nm in photon correlation spectroscopy. Release of all-trans-retinoic acid from the SAP nanoparticles was maintained over 5 weeks. For further study in vivo, we tested the cytotoxicity of SAP nanoparticles using an MTT assay, and cytotoxicity was augmented as the molar mass of stearic acid increased in human liver carcinoma HepG2 cells. Therefore, SAP nanoparticles might be a promising longterm delivery carrier for hydrophobic therapeutic molecules with the appropriate composition.Archives of Pharmacal Research 05/2010; 33(5):761-7. · 1.54 Impact Factor