Fabrication Of Functional Hyperbranched Conjugates And Their Applications In Nanomedicine

Gene and drug carriers are one of important research fields in nanomedicine. Hyperbranched polymer is widely used to overcome many deficiencies of the conventional carriers in this field due to its unique physical and chemical properties, such as low viscosity, many functional terminals, large internal cavity and high solubility. However, there are still a lot of issues, such as the contradiction between cytotoxicity and transfection efficiency, unsatisfied efficacy of individual gene therapy, and bacterial infection from low immunity. In this thesis, various functional hyperbranched conjugates have been constructed for improvement of treatment efficacy. The details and main conclusions are given as follows:1. Hyperbranched Glycoconjugated Polymer from Natural Small Molecule Kanamycin as a Safe and Efficient Gene VectorThe exploration of safe and efficient polycationic gene vectors from natural small molecules such as kanamycin is proposed. Cationic hyperbranched glycoconjugated polymer is synthesized by the Michael-addition polymerization of kanamycin and N’,N’-methylenebisacrylamide, and the resultant product is well characterized by Fourier transform infrared (FTIR),1H nuclear magnetic resonance (1H NMR),13C NMR, size exclusion chromatography-multiangle laser light scattering (SEC-MALLS) and ξ-potential analyses. The nitrogen content (7.3%) of this kanamycin-based hyperbranched glycoconjugated polymer is much lower than that (32.6%) of polyethylenimine (PEI) control. Moreover, this resultant polymer could be degraded in acidic conditions. Therefore, the hyperbranched glycoconjugated polymer shows low cytotoxicity, even lower than that of natural biomacromolecule chitosan. Due to the existence of various primary, secondary and tertiary amines in the polymer backbone, hyperbranched glycoconjugated polymer exhibits high buffering capacity and strong pDNA condensation ability. In vitro transfection shows that the luciferase expression of hyperbranched glycoconjugated polymer is about4.4×108RLU per mg protein, approximately33-fold greater than that of chitosan transfection. These results demonstrate that the construction of highly branched polycations from natural small molecules provides a new opportunity for developing safe and efficient gene vectors.2. A Controlled Drug Delivery System with Promotion of Gene Transfection and Anticancer Efficacy As an efficient method to improve the polycationic gene delivery, hydrophobic modification has received more and more attention. However, it is rarely reported that hydrophobic drugs are used to promote gene transfection. Considering that the target of hydrophobic chlorambucil is DNA strands, the hyperbranched polycations are modified with chlorambucil for both gene transfection and drug delivery. The hyperbranched glycoconjugated polymers are modified with chlorambucil, and its chemical structure is well analyzed by FTIR,1H NMR, Size Exclusion Chromatography (SEC) and biophysical properties. Since the polycations can transport drugs into the nucleus efficiently, the anticancer efficacy of drug conjugates is much better than that of free chlorambucil. The results from confocal laser scanning microscope and flow cytometry have verified the efficient cell internalization of polymeric carriers. Owning to the hydrophobic modification and the controlled release of chlorambucil from polycations, the hyperbranched glycoconjugated polymers with chlorambucil show high transfection efficiency than pure polymers.3. Multi-functional Hyperbranched Glycoconjugated Polymers Based on Natural AminoglycosidesThe multi-functional gene vectors with high transfection, low cytotoxicity, and good anti-tumor and anti-bacterial activities are prepared from natural aminoglycosides. Through the Michael-addition polymerization of gentamycin and N’,N’-methylenebisacrylamide, cationic hyperbranched glycoconjugated polymers are synthesized and their physical and chemical properties are well analyzed by FTIR,1H NMR,13C NMR, SEC, ξ-potential, and acid-base titration techniques. The cytotoxicity of these hyperbranched glycoconjugated polycations is low because of the hydrolysis of degradable glycosidic and amide linkages in acid conditions. Owing to the presence of various primary, secondary and tertiary amines in the polymers, hyperbranched glycoconjugated polymers show high buffering capacity and strong DNA condensation ability, resulting in the high transfection efficiency. In the meantime, due to the introduction of natural aminoglycosides into the polymeric backbone, the resultant hyperbranched glycoconjugated polymers inhibit the growth of cancer cells and bacteria efficiently. Combining the gene transfection, anti-tumor and anti-bacterial abilities together, the multi-functional hyperbranched glycoconjugated polymers based on natural aminoglycosides may play an important role in protecting cancer patients from bacterial infections.4. Sequential Drug Release for Synergistic Cancer Treatment and Immunity Promotion Effective drug sequential release can enhance the efficacy, so sequential delivery system is one of most important goals in cancer therapy. In this work, a sequential drug release for synergistic cancer treatment and immunity promotion has been constructed. Firstly, the hyperbranched glycidol (HPG) is synthesized by cationic ring-opening polymerization. Benefiting from the existence of many hydroxyl end-groups in HPG, the traditional Chinese medicine norcantharidin (NCTD) with anhydride can be readily conjugated onto polyols via ester linkages, forming the HPG-NCTD conjugates. Owing to the coordination between cisplatin (CDDP) and carboxyls in HPG-NCTD conjugates, the HPG-NCTD/CDDP complexes with nano scale are obtained. Both in vitro and in vivo evaluations show that the sequential release of CDDP and NCTD is achieved by combination of coordination connections and hydrolysable ester bonds. Correspondingly, a synergistic efficiency of cancer treatment and immunity promotion is realized. These experimental results confirm that the sequential release carriers based on coordination connections and degradable covalent bonds can be used to overcome the problems of leukopenia in cancer therapy, giving us a perspective in cancer treatment.