Chemo-enzymatic Synthesis Of Amphiphilic Random Polymeric Prodrugs, Their Self-assembly And Anticancer Activity

Polymer-drug conjugates and the assembled nanoparticles have attracted lots of attentions, since these systems offer numerous advantages compared with traditional small molecular drugs including reduced side effects, prolonged circulation time, controlled drug release and improved targeting ability.In this thesis, a chemo-enzymatic synthetic strategy was developed to prepare polymer-drug conjugates. Then the resulting drug-containing amphiphilic random copolymers were successfully self-assembled into micelles nanoparticles. The morphology of micelles, in vitro release behaviors, cell cytotoxicity and inhibiting effect of tumor growth to A549 cell derived xenograft tumor bearing Balb/C nude mice were further investigated. The thesis includes the following four parts.The enzymatic selective synthesis of multifunctional drug and sugar monomers was studied. Three kinds of polymerizable drug (cytarabine (Ara-C), fluorodeoxyuridine (FUDR) and gemcitabine) monomers were prepared by chemoselectivly or regioselectivly enzymatic transesterification in organic medium, and a series of drug-containing amphiphilic random copolymer were synthesised by radical copolymerization of drug monomers and sugar (glucose, galactose and lactose) monomers. Two monomethoxyl PEG methylacrylate with different PEG molecular weight (PEG350, PEG1000) were synthesized and chosen as comonomers. Then two kinds of gemcitabine-containing amphiphilic random copolymers with different gemcitabine content and PEG chain length were obtained by copolymerization of gemcitabine vinyl ester with the PEG comonomers.A series of polymeric micelles were fabricated by self-assembly of the resulting amphiphilic random coplymers in aqueous phase. Assembly ability of the glucose/galactose/lactose-functionalized and PEGylated drug-containing amphiphilic random copolymer was investigated, and assembly mechanism was preliminary discussed. Moreover, the influence of drug content, initial water content, standing time and temperature on morphology evolution of gemcitabine-containing nanoparticles was studied in detail. And the results indicated that the self-assembly nanoparticles of galactose/lactose-functionalized gemcitabine-containing could change from sphere to ringlike, then wormlike and lastly helical morphology.In vitro drug release behaviors and in vitro cytotoxicity of glucose or galactose-functionalized, cytarabine and FUDR-containing polymeric nanoparticles were studied. The results of drug release experiments showed both small molecular drugs could simultaneously be released from polymeric micelles in pH 7.4 PBS. In vitro cellular uptake and cytotoxicity experiments showed that micelles containing cytarabine and FUDR could be internalized by HepG2 hepatoma cells and significantly inhibit the growth of the cells. Meanwhile, the micelles containing galactose showed evident targeting function to HepG2 liver carcinoma cells, thus showed a better inhibition of tumor growth.Both in vitro and in vivo behaviors of PEGylated gemcitabine-containing micelles were investigated systematically. The results showed that gemcitabine molecules could be slowly released from both gemcitabine-containing micelles with different PEG molecular weight, and the release rate was relative to the pH value of incubation medium. Cellular uptake and in vitro cytotoxicity assays against Human cervical carcinoma (HeLa) cells, human lung cancerous cells (A549) and human hepatoma cells (HepG2) demonstrated that both PEGylated micelles could be effectively internalized by these cancer cells and induced cell apoptosis. In vivo experiment indicated that these micelles efficiently inhibited tumor growth when injected intravenously into A549 cell derived xenograft tumor bearing Balb/C nude mice.